SK15692002A3 - 3-Nitrogen-6,7-dioxygen steroids and uses related thereto - Google Patents

Abstract

A compound of formula (I) and pharmaceutical acceptable salts, solvates, stereoisomers and prodrugs thereof, in isolation or in mixture, wherein, independently at each occurrence: R<1> and R<2> are selected from hydrogen, oxygen so as to form nitro or oxime, amino, sulfate, and sulfonic acid, and organic groups having 1-30 carbons and optionally containing 1-6 heteroatoms selected from nitrogen, oxygen, phosphorous, silicon, and sulfur, where R<1> and R<2> may, together with the N to which they are both bonded, form a heterocyclic structure that may be part of an organic group having 1-30 carbons and optionally containing 1-6 heteroatoms selected from nitrogen, oxygen and silicon, and where R<1> may be a 2, or 3 atom chain to numeral 2 so that —N-R<1> - forms part of a fused bicyclic structure to ring A; R<3> and R<4> are selected from direct bonds to 6 and 7 respectively so as to form carbonyl groups, hydrogen, or a protecting group such that R<3> and/or R<4> is part of hydroxyl or carbonyl protecting group; numerals 1 through 17 each represent a carbon having substitution as described. The compounds may be formulated into pharmaceutical compositions, and used in the treatment and/or prevention of various conditions, including inflammation, asthma, an allergic disease, chronic obstructive pulmonary disease, atopic dermatitis, solid tumors, AIDS, ischemia, and cardiac arrhythmias.

Description

3-AMINO-6.7-DIOXYGENED STEROIDS AND USE / VWM £

Technical field

The present invention relates to 3-amino-6,7-dioxygenated steroids, compositions containing these steroids, and related therapeutic uses.

BACKGROUND OF THE INVENTION

Inflammatory response

Inflammation is the basic localized response of a host to invading organisms or tissue injury involving cells of the immune system. Classical signs of inflammation include redness (erythema), swelling (edema), pain, and increased heat (pyrene) at the site of the injury. The inflammatory response allows the body to specifically recognize and eliminate the invasive organism and / or repair tissue damage. Many of the acute changes at the site of inflammation can be directly or indirectly attributed to the massive intake of leukocytes (e.g., neutrophils, eosinophils, lymphocytes, monocytes) inherent to this response. Leukocyte infiltration and accumulation in tissue leads to activation and subsequent release of inflammatory mediators such as LTB ₄ , prostaglandins, TNF-α, IL-1β interleukins, IL-8, IL-5, IL-4, histamine, proteases and compounds with reactive oxygen.

Normal inflammation is a highly regulated process that is strictly controlled at several levels for each of the cell types involved in this response. For example, TNF-α pro-inflammatory cytokine expression is controlled at the level of gene expression, translation, post-translational modification, and release of the mature form from the cell membrane. Many of the proteins that are upregulated during inflammation are controlled by the NF-kappaB transcription factor. In some cases, pro-inflammatory responses are counteracted by endogenous anti-inflammatory mechanisms, such as IL-10 production. A normal inflammatory response is characterized by being temporary in nature, followed by a disease dispersal phase that restores the tissue's original condition. The disease dispersal phase is considered to include upregulation of anti-inflammatory mechanisms such as IL-10 as well as upregulation of pro-inflammatory processes.

Inflammatory diseases

Inflammatory diseases occur upon initiation of an inflammatory response that is inadequate or not resolved in the normal way, but rather persists and leads to a chronic inflammatory condition. The inflammatory disease may be systemic (e.g., lupus) or localized to particular tissues or organs and exhibits an extraordinary personnel and economic burden on the company. Examples of some of the most common and problematic inflammatory diseases include asthma, allergy, rheumatoid arthritis, inflammatory bowel disease, psoriasis, emphysema, colitis, graft versus host disease, contact dermatitis, and ischemic reperfusion injury. Other disease states, such as immunodeficiency diseases, are related to altered regulation of the chemokine / cytokine network and their receptors, which may alter viral replication and AIDS pathogenesis.

Many of the tissue, cellular, and biochemical processes that disrupt inflammatory disease have been elucidated, allowing the development of experimental models or tests to mimic the disease state. These in vitro and in vivo assays allow the selection and screening of compounds with a high probability of therapeutic efficacy in the respective inflammatory diseases. For example, the ability of a compound to inhibit allergen-induced inflammatory cell accumulation, such as eosinophils and lymphocytes, in irrigation fluid obtained from sensitized animals indicates anti-asthmatic activity. In particular, this model is useful for assessing the effects of compounds in the treatment of late-phase response and asthma-characteristic over-response in observable pulmonary inflammation. .

Asthma and allergy

Asthma and allergy are closely related to sufficient evidence from clinical trials showing a strong correlation between the severity of asthma and the degree of atopy (allergy). Sensitization to allergens is considered to be the most important risk factor for asthma in children and adults, where about 99% of asthma cases show atopy.

Allergy can be characterized by increased serum IgE (antibody). Repeated exposure to allergens in a process called sensitization is normally required to trigger atopy and a subsequent asthmatic and allergic response. B cells upon exposure to allergens provide antibodies that bind to the mast cell surface. Crosslinking of two antibodies by an antigen causes a series of reactions leading to degranulation and a transmission of a series of mediators that modulate the inflammatory response. Mediators that are released or formed during asthmatic and allergic responses include histamine, leukotrienes, prostaglandins, cytokines, and tryptase.

Asthma is characterized by excessive airway response, episodic bronchospasm and chronic pneumonia. Airway obstruction is reversible over time or as a result of response to drug therapies. Patients exhibiting normal air flow may be excessively reactive to a variety of natural stimuli, such as cold air, exercise, chemicals, and allergens. The most common case of initiating an asthmatic response is direct hypersensitivity to common allergens including weed pollen, grass pollen, various fungi, dust mites, cockroaches and pets. Symptoms of the disease include sore throat, wheezing, shortness of breath and cough. The incidence of asthma and mortality worldwide has increased and has doubled over the past 20 years despite the use of modern treatments.

Respiratory responses to allergen are complex and include an early asthmatic response (EAR) that peaks 20-30 min after challenge, can be characterized by bronchoconstriction and normally disappears within 1.5 to 2 h. Late asthmatic response (LAR) generally occurs 3 to 8 hours after initial exposure and involves both bronchoconstriction and the development of inflammation and edema in the lung tissue. This inflammation often becomes chronic with epithelial damage and lung infiltration by inflammatory cells such as eosinophils and neutrophils.

Current treatments for asthma

Glucocorticoids (steroids) are the most effective modality of long-term therapy for the treatment of asthma. For example, due to the presence of airway inflammation, even in mild asthma, inhaled steroids are used even at an early stage of drug therapy. Although steroids are effective anti-inflammatory agents, they are not very suitable for controlling acute onset asthma. Orally administered steroids have significant side effects and therefore their chronic use for asthma control is minimal. Combination therapy with orally delivered steroids is often used and can be divided into the following areas: anti-inflammatory drugs (for example, inhaled and oral steroids), bronchodilators (for example β ₂ -agonists, xanthines, anticholinergic agents) and mediator inhibitors (for example cromolines and leukotriene antagonists) ). In general, mild to severe asthma can be difficult to manage with the current drug arsenal. Drugs that are safe have only marginal efficacy, while effective medicines have unacceptable side effects requiring extensive patient monitoring. The drugs under development should meet the requirements for side effects (for example, vomiting-induced side effects typical of certain phosphodiesterase 4 inhibitors) and poor pharmacokinetic and metabolic parameters. There is a great need for therapeutic agents for the safe and effective treatment of inflammatory diseases such as asthma and allergy. The present invention provides these compositions and the related beneficial results described herein.

SUMMARY OF THE INVENTION

In one aspect, the invention provides compounds of Formula 1 and pharmaceutically acceptable salts, solvates, stereoisomers and prodrugs thereof, separately or in mixtures.

(d) in which, independently and in all cases:

R ¹ and R ² are selected from hydrogen, oxygen to form nitro or oxime, amino, -SO ₃ -R, and organic groups of 1 to 30 carbon atoms optionally containing 1 to 6 heteroatoms selected from atom nitrogen, oxygen, phosphorus, silicon and sulfur, where R ² may be a direct bond to atom number 3 or R ¹ and R ² together with the nitrogen atom to which they are both attached may form a heterocyclic structure which may be part of an organic group having 1 to 30 carbon atoms and may optionally contain 1 to 6 heteroatoms selected from nitrogen, oxygen and silicon, and R ¹ may be a chain of 2 to 3 atoms attached to atom number 2 so that -NR ¹ forms part of a fused bicyclic ring structure And,

R ³ and R ⁴ are selected from cases of direct bonds to atoms 6 and 7, respectively, in such a way as to obtain carbonyl groups, a hydrogen atom or a protecting group, such that R ³ and / or R ⁴ is part of a hydroxyl or carbonyl protecting group; marked with numbers 1 to 17 are all carbon atoms and the carbon atoms marked with numbers 1, 2, 4, 11, 12, 15, 16 and 17 can be independently substituted with (a) one of = O, = C (R ⁵ ) (R) ^5), = C = C ^{(R5) (R5)} -C ^{(R5) (R5)} (C ^{(R5) (R5))} n- and - (O (C ^(R5) ( R ⁵ ) is nO) -, wherein n is 1 to 6, or (b) two of the following groups, which are independently selected from -X,

-N (R ¹ ) (R ² ), -R ⁵ and -OR ⁶ , and the carbon atoms designated 5, 8, 9, 10, 13 and 14 may be independently substituted with one of -X, -R ⁵ , - N (R ¹ ) (R ² ) or -OR ⁶ , in addition to the groups -OR ³ and -OR ⁴ , each of carbon numbers 6 and 7 may each be independently substituted with one of -X, N- (R ¹ ) ( R ² ), -R ⁵ or -OR ⁶ , each of rings A, B, C, and D is independently fully saturated, partially saturated or fully unsaturated,

R ⁵ is each independently selected from hydrogen, halogen and C 1-3 organic, which may optionally contain at least one heteroatom selected from boron, halogen, nitrogen, oxygen, silicon and sulfur, wherein the two geminal R ⁵ groups may form a ring together with the carbon atom to which they both join,

R ⁶ is a hydrogen atom or a protecting group such that OR ⁶ is a protected hydroxyl group, wherein the vicinal -OR ⁶ groups may together form a cyclic structure that protects the vicinal hydroxyl groups and wherein the geminal -OR ⁶ groups may together form a cyclic structure that protects the carbonyl group and X represents fluoride, chloride, bromide and iodide.

In another aspect, the invention provides a pharmaceutical composition comprising a steroid compound as described above and a pharmaceutically acceptable carrier, excipient, or diluent.

In another aspect, the invention provides a method of treating inflammation comprising administering to a subject in need thereof a therapeutically effective amount of a steroid compound as described above.

In another aspect, the invention provides a method for the prophylactic treatment of inflammation comprising administering to a subject in need of such treatment a prophylactically effective amount of a steroid compound as described above.

In another aspect, the invention provides a method of treating asthma comprising administering to a subject in need thereof a therapeutically effective amount of a steroid compound as described above.

In another aspect, the invention provides a method of treating an allergic disease, including, but not limited to, dermal and ocular indications, comprising administering to a subject in need thereof a therapeutically effective amount of a steroid compound as described above.

In another aspect, the present invention provides a method of treating a chronic lung disease comprising administering to a subject in need of such treatment a therapeutically effective amount of a steroid compound as described above.

In another aspect, the invention provides a method of treating atopic dermatitis comprising administering to a subject in need of such treatment a therapeutically effective amount of a steroid compound as described above.

In another aspect, the invention provides a method of treating solid tumors comprising administering to a subject in need thereof a therapeutically effective amount of a steroid compound as described above.

In another aspect, the invention provides a method of treating AIDS comprising administering to a subject in need thereof a therapeutically effective amount of a steroid compound as described above.

In another aspect, the invention provides a method of treating ischemic reperfusion injury comprising administering to a subject in need of such treatment a therapeutically effective amount of a steroid compound as described above.

In another aspect, the present invention provides a method of treating cardiac arrhythmias comprising administering to a subject in need thereof a therapeutically effective amount of a steroid compound as described above.

These and related aspects of the invention are described in more detail below.

BRIEF DESCRIPTION OF THE DRAWINGS

Figures 1A and 1B show summary data on synthetic transformations that can be used to convert a 3-aminosteroid to a 3-nitrogensteroid of the invention.

Figures 2A, 2B, and 2C are a set of bar graphs showing the effect of compound 89 (dose-response, 4x daily, orally) on ovalbumin-induced accumulation of inflammatory cells in lung lavage fluid obtained from sensitized Brown Norway rats. Figure 2A shows the accumulation of eosinophils, Figure 2B shows the accumulation of neutrophils, Figure 2C shows the accumulation of lymphocytes.

Figures 3A, 3B and 3C provide a set of bar graphs showing the effect of compound 28 (dose-response, 4 times daily, orally) on ovalbumin-induced inflammatory cell accumulation in lung lavage fluid obtained from sensitized Brown Norway rats. Figure 2A shows the accumulation of eosinophils, Figure 2B shows the accumulation of neutrophils, Figure 2C shows the accumulation of lymphocytes.

Figure 4 is a graph showing the effect of test substances 28 and 89 administered orally once daily for 4 days prior to load on allergen-induced pulmonary resistance changes in sensitized guinea pigs.

Figure 5 is a graph showing the effect of test compounds 28 and 89 administered orally once daily for 4 days prior to load on allergen-induced changes in pulmonary elasticity in sensitized guinea pigs.

Figure 6 is a graph showing the duration of antibronchospastic effect of test compound 89 administered orally at 1 mg / kg once daily for 4 days prior to load on allergen-induced pulmonary resistance changes in sensitized guinea pigs.

Figure 7 is a graph showing the duration of antibronchospastic effect of test compound 89 administered orally at 1 mg / kg once daily for 4 days prior to load on allergen-induced pulmonary elasticity changes in sensitized guinea pigs.

A detailed description of the invention follows.

The present invention provides compounds, compositions and methods useful in the treatment and / or prevention of various disease states. For example, one aspect of the invention provides a method of treating and / or preventing an inflammatory disease. The method comprises administering to a subject in need of such treatment an effective amount of a compound of Formula 1 or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof, or an effective amount of a composition comprising a compound of Formula 1 or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof. .

Before describing the invention in more detail, certain terms are provided herein with the following definitions and certain conventions that are also used herein.

A. Definitions

As used herein, the following terms have the meaning indicated, unless the context clearly dictates otherwise.

Alkyl is a univalent, saturated or unsaturated, straight, branched, or cyclic, aliphatic (i.e., not aromatic) hydrocarbon group. In various embodiments, the alkyl group has 1 to 20 carbon atoms, i.e., a C1-C20 (or C1-C20) group, or a C1-C18, C1-C12, C1-C6 or C1-C4 group. In various embodiments, the alkyl group independently has zero branches (i.e., represents a straight chain), one branch, two branches or more than two branches, is saturated, unsaturated (where the unsaturated alkyl group may have one double bond, two double bonds, more than two double bonds and / or one triple bond, two triple bonds or more than two triple bonds), represents or comprises a cyclic structure, is acyclic. Examples of alkyl groups are C 1 alkyl (i.e. -CH ₃ (methyl)), C ₂ alkyl (i.e. -CH ₂ CH ₃ (ethyl), CH = CH ₂ (ethenyl) and -C = CH ( ethynyl) and C ₃ alkyl (e.g. -CH ₂ CH ₂ CH ₃ (n-propyl), -CH (CH ₃ ) ₂ (isopropyl), -CH = CH-CH ₃ (1-propenyl), -C = C-CH ₃ (1-propynyl), -CH ₃ -CH = CH ₂ (2-propenyl), -CH ₂ -C = CH (2-propynyl), -C (CH ₃ ) = CH ₂ (1-methyletenyl) and -CH (CH ₂ ) ₂ (cyclopropinyl)).

Aryl is a univalent aromatic hydrocarbon ring system. The ring system may be monocyclic or fused polycyclic (e.g., bicyclic, tricyclic, etc.). In various embodiments, the aryl ring is a C5-C10 or C5-C7 or C5-C6 ring, wherein the numbers of carbon atoms refer to the number of carbon atoms forming the ring system. The C6 ring system, i.e. the benzene ring, is a preferred aryl group. In various embodiments, the polycyclic ring is a bicyclic aryl group, wherein preferred bicyclic aryl groups are C8-C12 or C9-C10 groups. A naphthyl ring having 10 carbon atoms is a preferred polycyclic aryl group.

A heteroalkyl group is an alkyl group as defined herein in which at least one of the carbon atoms is replaced by a heteroatom. Preferred heteroatoms are nitrogen, oxygen, sulfur and halogen atoms. The heteroatom may, but usually does, have the same number of binding sites as the carbon atom. Accordingly, if a carbon atom is replaced by a heteroatom, the number of hydrogens attached to the heteroatom may be increased or decreased relative to the number of valence sites of the heteroatom. For example, if a carbon atom (4 valence) is replaced by a nitrogen atom (3 valence), then one of the hydrogen atoms attached in the first case to the replaced carbon atom must be omitted. Similarly, if a carbon atom is replaced by a halogen (one valence) then the three (all hydrogens) bound to the replaced carbon atom must be removed.

Heteroaryl group 'is a univalent aromatic system containing carbon atoms and at least one ring heteroatom. The heteroaryl group may, in various embodiments, have one heteroatom or one to two heteroatoms or one to three heteroatoms or one to four ring heteroatoms. The heteroaryl rings may be monocyclic or polycyclic, wherein the polycyclic ring may include a fused ring, a spiro compound ring, or a bridged ring. In one embodiment, the heteroaryl group is selected from a monocyclic and bicyclic system. Monocyclic heteroaryl rings may contain from about 5 to about 10 atoms (carbon and heteroatoms), preferably from 5 to 7 and most preferably from 5 to 6 ring atoms. The bicyclic heteroaryl rings may contain from about 8 to 12 members or 9 to 10 members in the ring. The heteroaryl ring may be unsubstituted or substituted. In one embodiment, the heteroaryl ring is unsubstituted. In another embodiment, the heteroaryl ring is substituted. Examples of heteroaryl rings include benzofuran, benzothiophene, furan, imidazole, indole, isothiazole, oxazole, piperazine, pyrosine, pyrazole, pyridazine, pyridine, pyrimidine, pyrrole, quinoline, thiazole, and thiophene rings.

Heteroatom ”is a halogen atom, nitrogen, oxygen, phosphorus, silicon or sulfur. Groups containing more than one heteroatom may contain different heteroatoms.

Hydrocarbons ”are chemical compounds consisting exclusively of hydrogen and carbon. Halocarbons are chemical compounds consisting exclusively of halogen and carbon and hydrohalocarbons ”are chemical compounds consisting exclusively of hydrogen, halogen and carbon.

Organic groups "and organic residues" are used synonymously and refer to stable structures having the number and type of atoms described.

The pharmaceutically acceptable salt &quot; and salts thereof &quot; in the case of the compounds of the present invention refer to acid addition salts and basic addition salts.

Acid addition salts refer to salts formed from the compounds of this invention and inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid and the like and / or organic acids such as acetic acid, propionic acid, glycolic acid pyruvic acid, oxalic acid, maleic acid, malic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid and the like.

Base addition salts refer to salts formed from the compounds of this invention and inorganic bases such as sodium, potassium, Utne, ammonium, calcium, magnesium, ferric, zinc, copper, manganese, aluminum salts and the like. Suitable salts include ammonium, potassium, sodium, calcium and magnesium salts and salts derived from pharmaceutically acceptable non-toxic bases such as primary, secondary and tertiary amines, substituted amines including natural substituted amines, cyclic amines and basic ion exchangers such as isopropylamine, trimethylamine, diethylamine, tripropylamine, ethanolamine, 2-dimethylaminoethanol, 2-diethylaminoethanol, trimetamine, dicyclohexylamine, lysine, arginine, histidine, caffeine, procaines, hydrabamine, choline, betaine, ethylenediamine, glucosamine, piperromine, piperidine, netobenzidine, teobromidine, teobromidine, teobromidine, teobromidine, teobromidine Similarly.

When any variable occurs more than once in any component or compound of Formula 1, its definition is always independent of the definition in any other occurrence. Combinations of substituents and / or variables are permissible only as long as these combinations result in stable compounds. The compounds useful in the methods and compositions of the invention, as well as the compounds of the invention, may have asymmetric centers and occur as racemates, racemic mixtures and as individual diastereomers or enantiomers, and all isomeric forms are included in the invention. The racemate or racemic mixture does not imply a 50:50 mixture of stereoisomers.

In another embodiment, the invention provides pharmaceutical compositions comprising a compound of Formula 1 as described above, in combination with a pharmaceutically acceptable carrier, diluent or excipient. These compositions can be used to treat inflammation or other conditions as described herein. These compositions may also be formulated as a medicament which may be used in the treatment of, for example, the treatment of inflammation.

These compositions are useful, for example, as testing standards, suitable bulk delivery formulations, or pharmaceutical formulations. A detectable amount of a compound of this invention is an amount that can be readily measured by standard assay methods well known and recognized by those of skill in the art. The detectable amounts of the compound of the invention will generally be from about 0.001% to about 100% by weight of the total composition. Inert carriers include any substance that does not decompose or react by other covalent reaction with a compound of formula 1. Examples of suitable inert carriers are water, aqueous buffers such as those commonly used in high performance liquid chromatography (HPLC), organic solvents such as is acetonitrile, ethyl acetate, hexane and the like; and pharmaceutically acceptable carriers.

Pharmaceutically acceptable carriers for therapeutic use are well known in the pharmaceutical art and are described, for example, in Remingtons Pharmaceutical Sciences, Mack Publishing Co. (A.R. Gennaro red., 1985). For example, sterile saline and phosphate buffered saline at physiological pH may be used. Preservatives, stabilizers, colorants and even flavors may be added to the pharmaceutical composition. For example, sodium benzoate, sorbic acid and phydroxybenzoic acid esters may be added as preservatives. Id. 1449. In addition, antioxidant and suspending agents may be used. Id.

The steroid compounds of the invention have at least four rings, commonly referred to as A, B, C, and D, as shown below, where ring A can be fused to another ring.

B. Compounds

The present invention provides compounds of Formula I and pharmaceutically acceptable salts, solvates, stereoisomers and prodrugs thereof, separately or in mixtures

(D in which independently and in all cases that

R ¹ and R ² are selected from hydrogen, oxygen to form nitro or oxime, amino, -SO 3 -R and organic groups of 1 to 30 carbon atoms optionally containing 1 to 6 heteroatoms selected from nitrogen oxygen, phosphorus, silicon and sulfur, where R ² may be a direct bond to atom number 3 or the R ¹ and R ² groups, together with the nitrogen atom to which they are both attached, may form a heterocyclic structure which may be part of an organic group with And may optionally contain 1 to 6 heteroatoms selected from nitrogen, oxygen and silicon, and R ¹ may be a chain of 2 to 3 atoms attached to atom number 2 so that -NR ¹ forms part of the condensed bicyclic structure of ring A , * 3 Λ

R and R are selected from cases of direct bonds to atoms 6 and 7, respectively, in such a way as to obtain carbonyl groups, a hydrogen atom or a protecting group, such that R ³ and / or R ⁴ is part of a hydroxyl or carbonyl protecting group; 1 to 17, all carbon atoms and carbon atoms designated 1, 2, 4, 11, 12, 15, 16 and 17 may be independently substituted with (a) one of = O, = C (R ⁵ ) (R ⁵ ) , = C = C (R ⁵ ) (R ⁵ ), -C (R ⁵ ) (R ⁵ ) (C (R ⁵ ) (R ⁵ )) n - and - (O (C (R ⁵ )) (R ⁵⁾ (i) nO) -, wherein n is 1 to 6, or (b) two of the following groups, which are independently selected from -X,

-N (R ¹ ) (R ² ), R ⁵ and -OR ⁶ , and the carbon atoms designated 5, 8, 9, 10, 13 and 14 may be independently substituted with one of -X, -R ⁵ , -N (R ¹ ) (R ² ) or -OR ⁶ , in addition to the groups -OR ³ and -OR ⁴ , each of carbon numbers 6 and 7 may be independently substituted with one of -X, -N (R ¹ ) (R ¹ ) ² ), -R ⁵ or -OR ⁶ , each of rings A, B, C and D is independently fully saturated, partially saturated or fully unsaturated,

R ⁵ is always independently selected from hydrogen, halogen and C1.30, which may optionally contain at least one heteroatom selected from boron, halogen, nitrogen, oxygen, silicon and sulfur, wherein the two geminal R ⁵ groups may form a ring together with the carbon atom to which they both join,

R ⁶ is a hydrogen atom or a protecting group such that OR ⁶ is a protected hydroxyl group, wherein the vicinal -OR ⁶ groups may together form a cyclic structure that protects the vicinal hydroxyl groups and wherein the geminal -OR ⁶ groups may together form a cyclic structure that protects the carbonyl group group and

X is fluoride, chloride, bromide and iodide.

In one aspect of the invention, R ¹ and R ² are selected from hydrogen and an organic group of 1 to 30 carbon atoms optionally containing 1 to 6 heteroatoms selected from nitrogen, oxygen, phosphorus, silicon and sulfur. R ² may optionally be a direct bond to atom number 3. In another aspect, R ¹ , R ² and the nitrogen atom to which both substituents are attached form a heterocyclic structure which may be part of an organic group having 1 to 30 carbon atoms and optionally contain 1 to 6 heteroatoms selected from nitrogen, oxygen and silicon. In another aspect, R ^{1 is a} diatomic or triatomic chain attached to atom number 2, so that -NR ¹ - forms part of the fused bicyclic structure with ring A and these 2 or 3 atoms are selected from carbon, nitrogen and oxygen when this results in a stable structure. In these other aspects of the invention, the organic group may have 1 to 20 carbon atoms, while in other optional embodiments the organic group has 1 to 10 carbon atoms.

In one preferred aspect of the invention, both R ¹ and R ² are hydrogen atoms. These steroids not only possess the desired biological activity, they can also serve as suitable precursor compounds for the preparation of other steroids of the invention in which R ¹ and / or R ² are not hydrogen atoms.

For example, the invention provides, in one embodiment, a compound of Formula 1 wherein R ¹ and R ² are hydrogen or protecting groups;

R and R are selected from cases of direct bonds to atoms 6 and 7, respectively, to form carbonyl groups, hydrogen atoms, such that R ³ and / or R ⁴ is part of the hydroxyl or carbonyl protecting groups and in addition to the -OR ³ and -OR ⁴ groups shown each of the 6 and 7 atoms substituted with a hydrogen atom, unless this is excluded, since -OR ³ or -OR ⁴ form a carbonyl group, carbon atoms 1, 2, 4, 11, 12, 15 and 16 are all substituted with two atoms hydrogen, carbon numbers 5, 8, 9 and 14 are all substituted with one hydrogen atom, carbon atom number is substituted with methyl, carbon atom 13 is substituted with methyl, unless it is part of an unsaturated bond, carbon atom number 17 is substituted ( (a) one of = 0, = C (R ⁵ ) (R ⁵ ), = C = C (R ⁵ ) (R ⁵ ), C (R ⁵ ) (R ⁵ ) (C (R ⁵ ) (R ⁵⁾ (R ( ⁵ )) n- and - (O (C (R ⁵ ) (R ⁵ )) n O) -, wherein n is 1 to 6, or (b) two of the following substituents independently selected: -X, -N (R ¹ ) (R ² ), -R ⁵ and -OR ⁶ , each of rings A, B, C and D independently being fully saturated, partially saturated or completely unsaturated, R ⁵ is each independently selected from hydrogen, X and C1.30 an organic radical which may optionally contain at least one heteroatom selected from boron, halogen, nitrogen, oxygen, silicon and sulfur, wherein the two geminal groups R ⁵ may together form the ring with the carbon atom to which they both attach, R ⁶ is a hydrogen atom or a protecting group, so -OR ⁶ is a protected hydroxyl group, where the vicinal -OR ⁶ groups may together form a cyclic structure that protects the vicinal hydroxyl groups and where geminal The groups -OR ⁶ may together form a cyclic structure that protects the carbonyl group and X represents fluoride, chloride, bromide and iodide.

In another embodiment, the invention provides a compound of formula 1 wherein R ¹ and R ² are hydrogen, R ³ and R ⁴ are selected from hydrogen and protecting groups such that R ³ and / or R ⁴ is part of a group hydroxyl protecting group, carbons number 1, 2, 4, 11, 12, 15 and 16 are all substituted by two hydrogen atoms, carbons number 5, 8, 9 and 14 are all substituted by one hydrogen atom, carbon number 10 is substituted by methyl, carbon number 13 is substituted with methyl, unless it is part of an unsaturated bond, carbon number 17 is substituted with (a) one of = C (R ⁵ ) (R ⁵ ) and = C = C (R ⁵ ) (R ⁵ ) or (b) the following two groups selected independently: -X, -N (R ¹ ) (R ² ) and -R ⁵ , each of rings A, B, C, and D independently being fully or partially saturated, R ⁵ is independently selected from the group consisting of hydrogen, X and C _1-30 hydrocarbons, halogen compounds and X represents fluoride, chloride, bromide and iodide.

In one further embodiment, the invention provides a compound of formula 1 wherein R ¹ and R ² are hydrogen atoms, R ³ and R ⁴ are selected from hydrogen atoms and protecting groups such that R ³ and / or R ⁴ is part of a group hydroxyl protecting group, carbon atoms number 1, 2, 4, 11, 12, 15 and 16 are all substituted by two hydrogen atoms, carbon atoms number 5, 8, 9 and 14 are all substituted by one hydrogen atom, carbon atom number 10 is substituted methyl, carbon 13 is substituted with methyl, unless it is part of an unsaturated bond, carbon 17 is substituted with one of = C (R ⁵ ) (R ⁵ ) and = C = C (R ⁵ ) ^(R5), and (b) two of the radicals R ^5, each of rings a, b, C and D is independently fully saturated or partially saturated, and R ⁵ are independently selected from hydrogen and hydrocarbon Cmo.

Specific compounds of the invention wherein R ¹ and R ² are hydrogen atoms include:

In another aspect, the invention provides steroids with a nitrogen substituent at the 3-position, wherein the nitrogen of the substituent is substituted with an organic group. For example, the present invention provides steroidal compounds wherein R ¹ is selected from -C (= O) -R ⁷ , -C (= O) NH-R ^7, and -SO 2 -R ⁷ wherein R ⁷ is selected from alkyl; a group, a heteroalkyl group, an aryl group, and a heteroaryl group. In a related embodiment, R ¹ is hydrogen and R ² is -CH ₂ -R ⁷ wherein R ^{7 is} selected from alkyl, heteroalkyl, aryl and heteroaryl. In one embodiment, R ^{7 is} selected from C 1-4 hydrocarbyl groups. In another embodiment, -C (= O) -R ⁷ comprises biotin. In another embodiment, R ^{7 is} selected from alkyl-substituted phenyl groups, halo-substituted phenyl groups, alkoxy-substituted phenyl groups, aryloxy-substituted phenyl groups, and nitrophenyl groups.

In another aspect, (R ¹ ) (R ² ) is an N-heterocyclic group, meaning that the carbon atom of the (R ¹ ) (R ² ) N- group may be part of a heterocyclic ring. Examples include

AAp / V »

Ä

In another aspect, at least one of R ¹ and R ^{2 is a} heterocyclic ring or a carbocyclic ring. A preferred heterocyclic ring is ^e

and the preferred carbocyclic ring is a benzene ring including a substituted benzene ring such as in the 3-methylphenyl group, the 4-hydroxyphenyl group and the 4-sulfonamidophenyl group.

In another aspect, R ¹ may be a diatomic or triatomic chain attached to atom number 2 such that NR ¹ - forms part of a bicyclic structure fused to ring A. This invention provides compounds of the formula shown below, wherein Z represents 2 or 3 atoms selected from carbon atoms , nitrogen and oxygen.

The ring comprising Z may be saturated or unsaturated.

t Í Y1

FROM·

I

N ·

As ⁷

Examples of these fused ring compounds include

In another aspect, R ¹ is hydrogen and R ² comprises a C 1-10 hydrocarbyl group.

In another aspect, R ¹ is hydrogen and R ² is heteroalkyl. Suitable heteroalkyl include, without limitation, (CVW alkyl) -N- (CVIOs alkylene) group where W is selected from the group of O and NH, HO- (C-io alkylene) group, and HO (alkylene CI.I ₀ N- (CI.I ₀ alkylene) group where W is selected from the group of O and NH.

In another aspect, both R ¹ and R ^{2 are} independently selected from hydrogen and C 1 -C 20 organic groups optionally containing 1 to 5 heteroatoms selected from nitrogen, oxygen, silicon, and sulfur.

In another aspect, each of R ¹ and R ^{2 is} independently selected from hydrogen, R ⁸ , R ⁹ , R ¹⁰ , R ¹¹ and R ¹² , wherein R ⁸ is selected from C 1-10 alkyl, C 1-10 heteroalkyl containing 1, 2 or 3 heteroatoms, a C 6-10 aryl group and a C 3-15 heteroaryl group containing 1, 2 or 3 heteroatoms, R ⁹ is selected from (R ⁸ ) r - (C _1-10 alkylene), (R ⁸⁾ 1 - (C 1-10 heteroalkylene) group containing 1, 2, 3 heteroatoms, (R ⁸ ) r (C 1-10 arylene) and (R ⁸ ) r - (C 3-15 heteroarylene) group containing 1, 2 or 3 heteroatoms R ¹⁰ is selected from the (R ⁹⁾ r (C ₁ .i ₀ alkylene) group, (R ⁹⁾ r (C 3-15 heteroarylene) having 1, 2 or 3 heteroatoms, R ¹¹ is selected from the (° R ¹⁾ r (Ci-io alkylene) group, ^(R1 °) r (Ci-io heteroalkylene) having 1, 2 or 3 heteroatoms, (R ¹⁰⁾ r (C 6 -C io arylene) group and a (R ¹⁰ ) r (C 3-15 heterorylene) group containing 1, 2, 3 heteroatoms, R ¹² is selected from the (R ¹¹⁾ r (C-io alkylene) group, a (R ¹¹⁾ r (C io heteroalkylene) having 1, 2 or 3 heteroatoms, (R ¹¹⁾ R - (C6-io arylene group), and (R ¹¹⁾ R (C3.is heteroarylene) having 1, 2 or 3 heteroatoms selected from the AR, the numbers 0, 1, 2, 3, 4 and 5, with the proviso that R ¹ and R ² may be attached to a common atom to form a ring with this common atom.

In another aspect, the invention provides steroid compounds of the structure shown above, wherein R ¹ and R ^{2 are} independently selected from hydrogen, R ⁸ , R ⁹ , R ¹⁰ , R ¹¹ and R ¹² , wherein R ⁸ is selected from alkyl; , heteroalkyl, aryl and heteroaryl, R ⁹ is selected from the (R ⁸⁾ ralkylénová group, ^(R8) r-heteroalkylene group, ^(R8) r-aryl-alkylene group, and (R ⁸⁾ r heteroarylene group, R ¹¹ is selected from the ^(R10) r-alkylene, (R ¹⁰⁾ rheteroalkylénová group, a (R ¹⁰⁾ rarylénová group, and ^(R10) r-heteroarylene, ^R12 is selected from the (R ¹¹⁾ ralkylénová group, ^(R11) r-heteroalkylene group, (R ¹¹⁾ rarylénová a group (R ¹¹⁾ L-heteroarylene Ar is chosen from the numbers 0, 1,2, 3, 4 and 5, with the proviso that the substituents R ¹ and R ² may be attached to a common atom in such a way that they form a ring with this common atom, R ³ and R ⁴ are selected from in the case of hydrogen and protecting groups such that R ³ and / or R ⁴ form part of a hydroxyl protecting group, carbon atoms 1, 2, 4, 11, 12, 15 and 16 are all substituted by two hydrogen atoms, carbon atoms number 5 , 8, 9 and 14 are all substituted with one hydrogen, carbon number 10 is substituted with methyl, carbon number 13 is substituted with methyl, unless it forms part of an unsaturated bond, carbon number 17 is substituted with one of = C (R ⁵ ) (R ⁵ ) a = C = C (R ⁵ ) (R ⁵ ) or (b) by two R ⁵ groups, each of rings A, B, C and D is independently fully saturated or partially saturated and R ⁵ is in each case selected from hydrogen and C1-10 hydrocarbon groups.

R ¹ and R ² can, for example, be selected from hydrogen, CH ₃ -, CH ₃ (CH ₂ ) 2 -, CH ₃ (CH ₂ ) 4 -, CH ₃ CO-, C ₅ H ₅ CO-, (CH ₃ ) ₂ -CHSO ₂ -, C ₆ H ₅ SO ₂ -, C ₆ H ₅ NHCO-,

CH ₃ (CH ₂ ) ₂ NHCO-, CH ₃ (CH ₂ ) 2 -NH (CH ₂ ) 2 -, (CH ₃ ) ₂ N (CH ₂ ) 2 -, HOCH 2 CH 2 -,

HOCH ₂ (CH ₂ ) 4-, HOCH ₂ -CH ₂ NHCH ₂ CH ₂ -, 3- (CH ₃ ) C ₆ H ₄ -, 4- (HO) C ₆ H ₄ -, 4- (H ₂ NSO ₂ ) C _b H ₄ - 4 - ((CH ₃ ) ₂ CH) C ₆ H ₄ -CH ₂ -, 2- (F) C ₆ H ₄ -CH ₂ -, 3- (CF ₃ ) C ₆ H ₄ -CH 2 -, 2- (CH ₃₎ C ₆ H ₄ CH ₂ -, 4- (CF ₃ O) C _e H ₄ -CH ₂ -, 3- _(C H ₅₎ C ₆ H ₄ -CH ₂ -, 3- (NO) C ₆ H ₄ -CH ₂ -

or R ¹ and R ² together with the nitrogen atom to which they are both attached may form a heterocyclic ring selected from the group consisting of:

Particular compounds of the invention R ² is not a hydrogen atom include wherein R ¹ is a hydrogen atom, however

One set of preferred compounds of the invention thus has a substituent R ¹ which is hydrogen and R ² is not hydrogen.

In the steroid compounds of the invention described herein, in one aspect R ³ and R ^{4 have} a hydrogen atom, i.e., the steroid has a hydroxyl substitution at each of the carbon atoms designated 6 and 7. In a related aspect, one or both of the hydroxyl groups are on carbons 6 and 7 in protected form, i.e., they are attached to a hydroxyl protecting group. Such protecting groups are well known in the art, for example, described in Green and Wuts Protective Groups in Organic Synthesis, John Wiley & Sons, New York, NY (1999). A suitable protecting group is a ketal group so that the present invention provides compounds of formula

IP ^ 13—

Í A

2 ^ \

-i /

rl

R ² i

As described above, the present invention provides steroidal compounds comprising compounds of defined stereochemistry. One such compound has the stereochemistry shown in the following structure for R ³ O- and R ⁴ O-.

* N 'í ²

2 ^ 1, 3.

* 10

1 * 4 5

- ^_8/14 15 '4' R ⁴

As also described above, the present invention provides the steroid salt forms of the invention, preferably pharmaceutically acceptable salts. In one embodiment, N (R ¹ ) (R ² ) is in the form of a salt. In other words, the group -N (R ¹ ) (R ² ) is protonated in such a way that the carbon atom carries a positive charge. In such cases, the steroid compound of the invention is an acid addition salt as defined herein. In one preferred aspect, the invention provides the hydrochloride salts of the steroid structures shown above. In another preferred aspect, the present invention provides acetate salts of the steroid structures described herein.

As also described above, the present invention provides prodrugs of certain compounds represented by Formula 1. In one aspect, the present invention is directed to a prodrug of any of the specific compounds represented by Formula 1. In another aspect, the present invention excludes prodrugs of specific compounds represented by Formula 1, thus, in one aspect, the present invention is directed to compounds of Formula I and pharmaceutically acceptable salts, solvates, stereoisomers, but not precursors thereof, separately or in admixture.

In the steroid compounds of the invention, as defined above, in a preferred embodiment, atom 17 is substituted with = C (R ⁵ ) (R ⁵ ) and R ⁵ is selected from hydrogen, halogen, C 1-6 alkyl, C 1-6 hydroxyalkyl and -CO ₂ - (C 1-6 alkyl). In another preferred embodiment, atom 17 is substituted with C 1-6 alkyl or C 1-6 alkyl. ₆ haloalkyl group or is substituted with -OR ⁶ or = 0, wherein R ⁶ is a hydrogen atom.

In the steroid compounds of the invention as defined herein, in a preferred embodiment at least one of the atoms 10 and 13 is substituted with a methyl group.

In the steroid compounds of the invention as defined herein, in a preferred embodiment, the numbers 1 to 16 represent all carbon atoms, wherein the carbon atoms of the numbers 1,2, 4, 11, 12, 15 and 16 may be independently substituted with (a) one of groups = O, = C (R ⁵ ) (R ⁵ ), = C = C (R ⁵ ) (R ⁵ ), -C (R ⁵ ) (R ⁵ ) (C (R ⁵ ) (R ⁵ )) n - and (O (C (R ⁵ ) (R ⁵ )) n O) -, wherein n is from 1 to about 6, or (b) two of the following groups independently selected from -X, -N (R ¹⁾ (R ² ), -R ⁵ and -OR ⁶ and the figure 17 represents a carbon atom substituted with (a) one of = C (R ^5a ) (R ^5a ) and = C = C (R ^5a ) (R ^5a ) and C (R ^5a ) (R ^5a ) C (R ^5a ) n -, wherein n is from 1 to about 6, or (b) two of the following groups are independently selected from the following: -X, -N (R ¹ ) (R ² ) and -R ^5a , wherein R ^5a is in each case independently selected from hydrogen, halogen and C1.30 organic radical, which may optionally contain at least one heteroatom selected from cases and boron, halogen, nitrogen, silicon and sulfur, wherein the two geminal groups R ⁵ may form a ring together with the carbon atom to which they both attach. Optionally in each case can be selected independently from R ^{5 a} C 1-30 hydrocarbon group, C 1-30 halocarbon group, C4.30 hydrohalogénkarbónovej group, a hydrogen atom and a halogen atom. In an alternative embodiment, in all cases, R ^{5a can be} independently selected from the group of C 1-10 hydrocarbon group, C 1-10 halogencarbone group, C 1-4 hydrohalocarbon group, hydrogen atom and halogen atom. Optionally, the present invention provides in each of these embodiments other embodiments wherein R ¹ and R ² are selected from hydrogen, oxygen in such a manner as to form nitro, oxime, amino, -SO 3 -R, and organic groups having 1 to 30 carbon atoms optionally containing 1 to 6 heteroatoms selected from oxygen, phosphorus, silicon, sulfur, where R ² may be a direct bond to atom 3 or R ¹ and R ² may form together with the nitrogen atom to which they both join, the heterocyclic structure, which may be part of an organic group having 1 to 30 carbon atoms and optionally may contain 1 to 6 heteroatoms selected from oxygen and silicon, or R ¹ may be a diatomic or triatomic chain attached to atom number 2 such that the -NR ¹ - forms part of the fused bicyclic structure with ring A. Alternatively, the invention provides in k in each of these embodiments, further embodiments wherein all carbon numbers 1,2,4,11, 12, 15 and 16 are substituted with two hydrogen atoms, unless the carbon atom is part of an unsaturated bond, carbon atoms number 5, 8, 9 and 14 are all substituted with one hydrogen atom, unless this carbon atom is part of an unsaturated bond, carbon atom 10 is substituted with methyl, and carbon number 13 is substituted with methyl, unless it is part of an unsaturated bond. In each of these embodiments, the present invention optionally provides another embodiment wherein the carbon atoms 1, 2, 4, 11, 12, 15 and 16 are all substituted with two hydrogen atoms, carbon atoms number 5, 8, 9 and 14 are all substituted. one hydrogen atom, carbon number 10 is substituted with methyl, and carbon number 13 is substituted with methyl, unless it is part of an unsaturated bond.

In the steroid compounds of the invention as defined herein, in a preferred embodiment, both R and R are both hydrogen and / or both R ³ and R ⁴ are hydrogen and / or carbon number 17 is substituted with one of the following: groups: C (R ^5a ) (R ^5a ), = C = C (R ^5a ) (R ^5a ),

C (R ^5a ) (R ^5a ) (C (R ^5a ) (C (R ^5a ) (R ^5a )) _n , wherein n is from 1 to about 6, or (b) two of the following groups independently selected from: of cases -X, -N (R ¹ ) (R ² ) and -R ^5a , wherein R ^5a is in each case independently selected from hydrogen, halogen, and a C 1-10 organic radical which may optionally contain at least one heteroatom selected from from boron, halogen, nitrogen, silicon and sulfur atoms, wherein the two geminal R ⁵ groups together with the carbon atom to which they both are attached may form a ring.

In the steroids of the present invention, unless otherwise described, each of rings A, B, C and D is independently fully saturated, partially saturated or completely unsaturated. This means that the hydrogen atoms attached to any of the carbon atoms at positions 1 to 17 can be omitted in such a way as to allow unsaturation within the ring A, B, C and / or D. For example, if carbon atoms number 5, 8, 9 and 14 disclose as substituted with one hydrogen atom, and it is also disclosed that each of rings A, B, C and D is independently fully saturated, partially saturated or fully unsaturated, then any one or more of the hydrogen atoms attached to carbon atoms may be 5, 8, 9 and 14 omitted to allow carbon saturation.

The compounds of this invention are contemplated as pharmaceutical compositions. Preferably, the molecular weight of the compound of the invention is relatively small, i.e. less than about 5000 g / mol, usually less than 4000 g / mol, more usually less than 3000 g / mol, even more usually less than 2000 g / mol, and more usually less than 1000 g / mol, wherein the minimum molecular weight of the compound of the invention is about 300 g / mol, and each of these typical ranges is a particular embodiment of the invention.

The steroid compounds of the invention include pharmaceutically acceptable salts, solvates, stereoisomers and precursors of steroid structures with a nitrogen group at the 3-position, dioxygenated at the 6, 7-positions, separately or in admixture with each other.

The steroid compounds of the invention may exist and usually exist as solids, including crystalline solids, which can be crystallized from conventional solvents such as ethanol, Ν, Ν-dimethylformamide, water or the like. The crystallization process may provide different polymorphic structures depending on the crystallization conditions. Typically, a thermodynamically stable form of the steroid compound of the invention is preferred for commercial scale production and is the preferred form of the compound.

Crystallization often provides a solvate of the steroid compound of the general structure shown above. The term solvate, as used herein, refers to an aggregate comprising one or more nitrogen-substituted 3-position steroid compounds dioxygenated at positions 6, 7 of the invention with one or more solvent molecules. The solvent may be water, in which case the solvate is a hydrate. Alternatively, the solvent may be an organic solvent. The compounds of the invention may therefore exist as a hydrate including monohydrate, dihydrate, hemihydrate, sesquihydrate, trihydrate, tetrahydrate and the like, as well as other corresponding solvated forms. The steroid compounds may be true solvates, while in other cases the steroid may only retain the accidentally present water or may be a mixture with water and some randomly present solvent.

The term "pharmaceutically acceptable solvate" as used herein refers to a solvate that retains the biological activity and properties of biologically active steroid compounds having a nitrogen group at the 3-position, dioxygenated at the 6, 7-positions, of the present invention. Examples of pharmaceutically acceptable solvates include, but are not limited to, water, isopropanol, ethanol, methanol, dimethylsulfoxide, ethyl acetate, acetic acid, and ethanolamine. One skilled in the art will appreciate that the solvated forms are equivalent to the unsolvated forms and are intended to be included within the scope of this invention. An example of a reference describing solvates is P.A. Sykes, Guidebook to Mechanism in Organic Chemistry, 6th Edition (1986, John Wiley &amp; Sons, N.Y.).

The compounds of the invention may exist as individual stereoisomers, racemates and / or mixtures of enantiomers and / or diastereomers. All of these individual stereoisomers, racemates and mixtures thereof are contemplated within the scope of this invention. In one preferred aspect, the compounds of the invention are used in optically pure form.

The term "pharmaceutically acceptable prodrug" is intended to mean a compound that can be converted under physiological conditions or by solvolysis to a biologically active steroid compound having a nitrogen group at the 3-position, dioxygenated at the 6, 7-positions, as described above. Thus, the term prodrug refers to a metabolic prodrug of a steroid compound of the invention that is pharmaceutically acceptable. The prodrug may be inactive when administered to a subject, but is converted in vivo to an active steroid compound having a nitrogen group at the 3-position, dioxygenated at the 6, 7-positions of the invention. Drug prodrugs are usually rapidly transferred in vivo to give the parent compound of the above formulas, for example, by hydrolysis in blood.

A discussion of drug precursors is provided by T. Higuchi and V. Stella Prodrugs as Novel Delivery Systems, Volume 14 of the ACS Symposium Šerieš and Bioreversible Carriers in Drug Design, editors of Edward B. Roche, American Pharmaceutical Association and Pergamon Press, 1987, and both are incorporated herein by reference. A conventional drug prodrug is a derivative of the steroid compounds of the invention having chemically and metabolically cleavable groups and becoming solvolysis or under physiological conditions the compounds of the invention that are pharmaceutically active in vivo. The drug prodrug form often offers advantages in solubility, tissue compatibility or delayed release in mammalian organisms (see H. Bundgard, Design of Products, pp. 7-9, 2127).

24, Elsevier, Amsterdam 1985). A preferred prodrug is a compound having a substitution at the 3 steroid nitrogen atom of the invention, wherein the substituent is cleaved in vivo to yield a pharmaceutically active compound.

The steroids of the invention having a nitrogen substitution at the C3 atom and an oxygen substitution at the 6 and 7 positions have unexpected potency enhancing properties for these compounds. For example, the steroid of the present invention exhibits excellent metabolic stability in human liver S9 fractions. For example, 100% of compounds 28, 89, 139 and 143 remain unchanged after 15 and even 30 min incubation with human S9 fractions. An unexpected finding was that nitrogen substitution groups on C3 carbon significantly increased glucuronidation of molecules in blood plasma. In addition, the steroids of the invention having nitrogen-containing substituents on C3, such as compounds 28, 89 and 83, are highly soluble in aqueous solution and exhibit a solubility greater than 100 mg / ml in water. Furthermore, the efficacy and pharmacokinetic profile of the steroids of the present invention with nitrogen substitution groups at C3 are highly suitable for therapeutic use. Doses <1.0 mg / kg once daily reproducible demonstrate significant anti-inflammatory efficacy in in vivo inflammatory models. In rats, compounds 28 and 89 have an average half-life of 7.5 h and an oral bioavailability of approximately 100%, whereas in monkeys the half-life averages 15 h and an oral bioavailability of 25 to 30%. The maximum blood plasma concentration of both species is predictable and linear.

C. Preparation of Compounds

The compounds of the invention may be prepared by methods employing steps known to those skilled in the art or analogous thereto. General methods for carrying out reactions of steroid compounds can be found in Steroid Reactions, C. Djerassi, ed. Holden Day, San Francisco, CA. 1963 and references cited therein. General methods of preparation can be found in Comprehensive Organic Transformations ”, R. C. Larock, VCH Publishers, New York, N.Y., 1989, and references cited therein. Further literature references useful in the preparation of the compounds of this invention are T. Reichstein, C. H. Meystre, Helv. Chim. Acta, 22, 728 (1932); H. Westmijze; H. Kleyn; P. Vermeer. L.A. van Dijck, Tet. Lett., 21, 2665 (1980), K. Prezewowsky, R. Wiechert, U.S. Pat. 3,682,983, P. Kaspar, H. Witzel,

J. Steroid Biochem., 23, 259 (1985), W.G. Dauben, T. Brookhart, J. Am. Chem.

Soc., 103, 237 (1981), A.J. Manson et al., J. Med. Chem., 6, 1 (1963), R.O. Clinton et al., J. Am. Chem. Soc., 83, 1478 (1961) and J.A. Zderie, et al., Chem. and Ind.,

1960. 1625.

In one preferred embodiment, steroids polyoxygenated at C3, C6, C7 and C17 are used as starting materials or intermediates. Methods for introducing oxygen at the C6 and C7 positions into commercially available starting materials are described in U.S. Pat. This patent also discloses many methods by which substitution and preferred stereochemical configuration can be ensured at positions C1, C2, C4, C5, C8, C9, C10, C11, C12, C13, C14, C15, C16 and C17. In the present invention, oxygen may be present in the C6 and C7 substituents in the form of hydroxyl groups or protected hydroxyl groups. The 6- and 7-hydroxyl groups may be protected individually or together may form part of a ring. Suitable protecting groups are listed in Greene and Wuts Protective Groups in Organic Synthesis, John Wiley & Sons, New York, N.Y. (1999).

Referring to Scheme A, the ketones of compound 2 or analogous compounds can be alkylated with a plurality of alkylation groups to give steroids of the invention having, but not limited to, alkyl, cycloalkyl, aryl and heteroaryl substituent groups. For example, alkylation of 17-ketone 2 with an acetylene anion provides the 17α-ethynyl-17β-hydroxyl intermediate 3. Inverting the stereochemistry of the C 17 substituents can be accomplished by first forming the methyl sulfonate followed by treatment with silver nitrate in tetrahydrofuran and water. Dehydration of compound 3 with phosphorus oxychloride in 2,4-lutidine gives compound 4. Tetrabutylammonium fluoride in tetrahydrofuran removes the tert-butyldimethylsilyl protecting group from the 3-hydroxyl group to give compound 5. Treatment of the 3α-hydroxyl compound 5 with ZnN ₆ -2py, diisopropylphosphine and diisophenyl phosphine ) in toluene gives the 3β-azide compound 6. ZnN6-2py is prepared by reaction with zinc nitrate and sodium nitride followed by pyridine treatment according to MC Viaud and P. Rollina in Synthesis 1990, 130. Reduction of the azide with lithium aluminum hydride in diethyl ether provides amine 7. Treatment hydrochloric acid in tetrahydrofuran and water removes the acetonide group to give the ammonium chloride salt 8.

i) KCCH, ii) POCl ₃ , 2,4-lutidine, iii) Bu ₄ NF, THF, iv) ZnN ₆ -2py, Ph ₃ P, DIAD, toluene, iii) L 1 AlH 4, Et ₂ O, iv) HCl, water, MeCN.

According to Scheme B, an alene-functional steroid of the present invention can be prepared from intermediates analogous to compound 3. An example is the reaction of compound 3 with lithium aluminum hydride and aluminum chloride in tetrahydrofuran to give alene 9. Tetrabutylammonium fluoride in tetrahydrofuran removes the protecting group from 3-hydroxy recovery of compound 10. Treatment of the 3-hydroxy group of compound 10 with ZnN ₆ -2py, triphenylphosphine and DIAD in toluene affords the 3β-azide compound 11. Reduction of azide 11 with lithium aluminum hydride in diethyl ether gives amine 12. Treatment with hydrochloric acid in tetrahydrofuran and water eliminates acetone 13th

Scheme B

i) LiAlH ₄ , AlCl 3, THF, ii) B 114 NF, THF, iii) ZnN ₆ 2py, Ph ₃ P, DIAD, toluene, iv) LiAlH ₄ , Et ₃ O, v) HCl, water, MeCN.

According to Scheme C, alkynyl-functional compounds of the invention can be prepared from alene intermediates. An example is treatment of compound 9 with n-butyllithium in tetrahydrofuran to give 17β-ethynyl compound 14. Tetrabutylammonium fluoride in tetrahydrofuran removes the protecting group from the 3-hydroxy group to give compound 15. Treatment of the 3α-hydroxyl compound 15 with ZnN ₆ -2py, triphenylphosphine and DIAD in toluene reduction of azide 16 with lithium aluminum hydride in diethyl ether affords amine 17. Treatment with hydrochloric acid in tetrahydrofuran and water removes the acetonide group to give the ammonium chloride salt 18.

i) n-BuLi, THF, ii) Bu ₄ NF, THF, iii) ZnN ₆ -2py, Ph ₃ P, DIAD, toluene, iv) LiAlH ₄ , Et ₂ O, v)

HCl, water, MeCN.

According to Scheme D, steroids of the present invention with an alkenyl functional group can be prepared from alkyne intermediates. An example is the controlled hydrogenation of compound 14 using palladium on calcium carbonate as a catalyst to obtain alkene 19. Tetrabutylammonium fluoride in tetrahydrofuran removes the protecting group from the 3-hydroxyl group to give compound 20. Treatment of the 3α-hydroxyl compound 20 with ZnN ₆ -2py, triphenylphosphine and DIAD in toluene provides the 30-azide compound 21. Reduction of azide 21 with lithium aluminum hydride in diethyl ether affords amine 22. Treatment with hydrochloric acid in tetrahydrofuran and water removes the acetonide group to give the ammonium chloride salt 23.

Scheme D

i) H ₂ , Pd-CaCO ₃ , ii) Bu ₄ NF, THF, iii) ZnN ₆ -2py, Ph ₃ P, DIAD, toluene, iv) LiAlH ₄ , Et ₂ O, v) HCl, water, MeCN.

Compound 2 can be used in a variety of olefinance reactions, including Wittig-type reactions to give a compound of the invention with an exocyclic olefinic group on C17. For example, as shown in Scheme E, compound 2 can be treated with ethyl triphenylphosphonium bromide and potassium (tert-butoxide) (KO'Bu) to give compound 24 wherein R ¹ is methyl and R ² is hydrogen. Tetrabutylammonium fluoride in tetrahydrofuran removes the protecting group from the 3-hydroxyl group to give compound 25. Treatment of the 3α-hydroxyl compound 25 with ZnN6-2py, triphenylphosphine and DIAD in toluene affords the 3β-azide compound 26. Reduction of azide 26 with lithium aminoethyl hydride provides 27 hydrochloric acid in tetrahydrofuran and water removes the acetonide group to give the ammonium chloride salt in which R ¹ is a methyl group and R ² is a hydrogen atom.

Analogously to the preparation depicted in Scheme E, ketones such as compound 2 can be reacted with other Wittig-type reagents such as, but not limited to, methyl, propyl, butyl, pentyl or hexyltriphenylphosphonium bromide to provide the steroids of the invention analogous to compound 28 wherein R ² is hydrogen and R ¹ is hydrogen, ethyl, propyl, butyl or pentyl.

i) EtPPh ₃ Br, KO ¹ Bu, toluene, ii) Bu ₄ NF, THF, iii) ZnN ₆ -2py, Ph ₃ P, DIAD, toluene, iv)

LiAlH ₄ , Et ₂ O, v) HCl, water, MeCN.

The steroids of the present invention may contain exocyclic double bonds of E and / or Z geometry. For example, as shown in Scheme F, Zolefin 24 in cyclohexane may be treated with ultraviolet light in the presence of diphenyldisulfide resulting in isomerization to E-olefin 29. Tetrabutylammonium fluoride in tetrahydrofuran removes treatment of 3α-hydroxyl compound 30 with ZnN ₆ -2py, triphenylphosphine and DIAD in toluene affords the 3β-azide compound 31. Reduction of azide 31 with lithium aluminum hydride in diethyl ether provides amine 32. Treatment with hydrochloric acid in tetrahydrofuran and tetrahydrofuran and tetrahydrofuran. water removes the acetonide group to form the ammonium chloride salt 33.

Scheme F

i) (PhS) ₂ , UV light, cyclohexane, ii) Bu ₄ NF, THF, iii) ZnN 6-2py, PhaP, DIAD, toluene, iv) LiAlH ₄ , Et ₂ O, v) HCl, water, MeCN.

Many side-chain functional steroids of the invention can be prepared by methods such as Lewis acid-supported attachment to aldehydes and Michael acceptors. For example, as shown in Scheme G, 24 can be reacted with methyl propionate in the presence of diethylaluminum chloride to give compound 34. Double bonds can be hydrogenated in the presence of a catalyst such as platinum to give compound 35. Tetrabutylammonium fluoride in tetrahydrofuran removes the protecting group from 3hydroxyl Treatment of the 3α-hydroxyl compound 36 with ZnN6-2py, triphenylphosphine and DIAD in toluene affords the 3β-azide compound 37. Hydrogenation of the azide 37 in the presence of a palladium catalyst affords the amine 38. Treatment with hydrochloric acid in tetrahydrofuran and water removes the acetonide group after formation of the ammonium chloride salt 39.

Scheme G

i) HCCCO ₂ Me, Et ₂ AlCl, ii) H ₂ , Pt, iii) Bu ₄ NF, THF, iv) ZnN ₆ -2py, Ph ₃ P, DIAD, toluene, v) H ₂ , Pd, EtOAc, vi 80% acetic acid.

Many secondary and tertiary amino compounds of the invention can be prepared from the 3-aminosteroid prepared according to any one of Schemes A through G above. Figures 1A and 1B illustrate several preparation methods that can be used to obtain the 3-amino compounds of the invention. For example, reductive amination may be used to link primary (see Figure 1A) and secondary (see Figure 1B) amines with aldehydes (RC (= O) H) and ketones (RC (= O) R '). 1A or 1B, the compounds may react with two aldehyde groups, i.e. dialdehydes of the formula HC (= O) -RC (= O) H with 3-aminosteroids to give steroids with heterocyclic structures at the 3-position. In addition (or alternatively) use reductive amination methods to attach 3-ketosteroids to heterocyclic secondary amines. Through these methods, the present invention provides compounds wherein R ¹ and R ² together with the nitrogen atom to which they are both attached may form a heterocyclic structure which may be part of an organic group having 1 to 30 carbon atoms, optionally containing 1 to 30 carbon atoms. 6 heteroatoms selected from nitrogen, oxygen and silicon atoms. Commercial sources and references to the chemical literature provide those of skill in the art with access to a variety of aldehydes (including dialdehydes) and ketones that can be used to prepare the steroid compounds of the invention. Methods of reductive amination are described, for example, in Synthesis, 1975, 135, J. Am. Chem. Soc., 93, 2897 (1971), M. Freifelder in Catalytic Hydrogenation in Organic Synthesis, L. Wiley & Sons 1978, Chapter 10, Russ. Chem. Rev., 49, 14 (1980) and references cited therein. See also J. Chem. Soc. Perkin Trans, 1, 2527 (1998) and Synlett, 1999, 1781, as well as the references cited herein.

Primary (see Figure 1A) and secondary (see Figure 1B) amines can be coupled with aryl compounds (ArX) to form a number of aryl-substituted amino compounds of the invention. Commercial sources and references to the chemical literature provide those of ordinary skill in the art with access to many aryl compounds that can be used to prepare steroid compounds of the invention. Examples of methods for amination of aminoaryl compounds can be found in J. Org. Chem., 65, 1158 (2000) and reviewed in Angew. Chem. Int. Ed., 37, 2046 (1998) and references cited therein.

Methods of reacting primary (see Figure 1) and secondary (Figure 1B) amines with acyl chlorides (RC, (= O) Cl) and sulfonyl chlorides (RSO ₂ Cl) to form amides and sulfonamides of the present invention are well known to those skilled in the art. organic chemistry in connection with other amino compounds and the same methods can be used for the amino compounds of the invention. Commercial sources and references to the chemical literature provide one of ordinary skill in the art with access to many of the acyl chlorides and sulfonyl chlorides that can be used to prepare the steroid compounds of the invention.

Methods of reacting primary (see Figure 1A) and secondary (see Figure 1B) amines with isocyanates (RN = C = O) and isothiocyanates (RN = C = S) to form ureas and thioureas, respectively, are also well known to one of skill in the art. organic chemistry in connection with other amino compounds and these methods can be used for the amino compounds of the invention. Commercial sources and references to the chemical literature provide those of ordinary skill in the art with access to a variety of isocyanates and isothiocyanates that can be used to prepare the steroid compounds of the invention. Review article Russ. Chem. Rev., 54, 249 (1985) and references cited herein describe examples of many substituted ureas and thioureas that may be included in the present invention.

Thus, using appropriately selected aldehydes, ketones, aryl compounds, acyl chlorides, sulfonyl chlorides, isocyanates and / or isothiocyanates, one skilled in the art can prepare steroid compounds wherein R ¹ and R ^{2 are} selected from hydrogen and organic groups. having 1 to 30 carbon atoms optionally containing 1 to 6 heteroatoms selected from nitrogen, oxygen, phosphorus, silicon and sulfur atoms.

The steroids of the invention may have fused heterocyclic rings such as, but not limited to, pyrazole, isoxazole and pyrimidine rings. As shown in Scheme H, compound 43 is an example of a fused pyrazole of the present invention for which the preparation of starting material 40 is described in U.S. Patent 6,046,185. Treatment of compound 40 with ethyl formate in pyridine in the presence of sodium methoxide affords the hydroxymethylene intermediate 41. with hydrazine hydrate in ethanol affords the pyrazole compound 42, which upon treatment with tetrabutylammonium fluoride in tetrahydrofuran provides the compound

43rd

Scheme H

i) EtO ₂ CH, NaOMe, pyridine, ii) N ₂ H ₄ , EtOH, iii) Bu ₄ NF ₃ , THF.

As shown in Scheme I, intermediates such as compound 41 can be converted to isoxazoles, exemplified by compound 44. Treatment of hydroxymethylene intermediate 41 with ammonium hydroxide in pyridine with further deprotection of hydroxyls at positions 6 and 7 with tetrabutylammonium fluoride in tetrahydrofuran affords isoxazole

44th

Scheme I

i) HONH.HCl, pyridine; ii) Bu ₄ NF, THF.

As shown in Scheme J, intermediates such as compound 41 can be converted to pyrimidines, exemplified by compound 44a. Treatment of the hydroxymethylene intermediate 41 with benzamidine hydrochloride and potassium hydroxide in ethanol followed by deprotection of the hydroxyls at positions 6 and 7 with tetrabutylammonium fluoride in tetrahydrofuran affords pyrimidine 44a.

Scheme J

i) benzamidine hydrochloride, KOH, EtOH, ii) BiuNF, THF.

By continuing through compounds with a ketone substitution at the 3-position and a substitution of = CHOH at the carbon 2 position, the present invention thus provides access to many compounds in which R ¹ can be a diatomic or triatomic chain attached to carbon 2 so that -NR ¹ - forms part of a fused bicyclic structures with circle A.

The reaction of 3-ketosteroids with hydroxylamine and pyridine can be used to obtain the steroid oximes of the invention. The steroidal oxime has R ² as a direct bond to carbon 3, thereby providing a double bond between carbon 3 and nitrogen and R ¹ is a hydroxyl group. Primary amines can be oxidized to nitro compounds, for example with dimethyldioxirane. Thus, R ¹ and R ² may be oxygen atoms. Methods for forming a nitro group are described in J. Org. Chem., 54, 5783 (1989). Reaction of 3-ketones with dimethylhydrazine affords the Ν, Ν-dimethylhydrazone steroids of the invention wherein R ² is a direct bond to atom 3 and R ¹ is NMe ₂ . Hydrazine treatment of dimethylhydrazone steroids provides the hydrazone steroids of the invention. A description of methods for the formation of Ν, Ν-dimethylhydrazones and hydrazones can be found in J. Org. Chem., 31, 677 (1966). The primary amines can also be reacted in a manner known in the art with sulfonic acids / sulfuric acid and esters to give sulfamate compounds, i.e., steroids in which atom 3 is bonded to -N-SO 3 -R and R is a hydrogen atom or an organic group having 1 to 30 carbon atoms optionally containing 1 to 6 heteroatoms selected from nitrogen, oxygen, phosphorus, silicon and sulfur atoms.

D. Pharmaceutical preparations

The present invention provides a pharmaceutical or veterinary composition (hereinafter referred to as a pharmaceutical composition) comprising a compound of Formula 1 as described above in admixture with a pharmaceutically acceptable carrier. The present invention further provides a composition, preferably a pharmaceutical composition, comprising an effective amount of a compound described above together with a pharmaceutically acceptable carrier.

The pharmaceutical compositions of the invention may be in any form that allows administration of the composition to a patient. For example, the preparation may be in the form of a solid, liquid or gaseous (aerosol). Conventional routes of administration include, without limitation, oral, topical, parenteral, sublingual, rectal, vaginal, ocular, and intranasal administration. The term parenterally as used herein includes subcutaneous injections, intravenous, intramuscular, intrasternal injections or infusion routes. The pharmaceutical composition of the invention is formulated in such a way that the active ingredients it contains are bioavailable to the patient when the composition is administered. The formulations to be administered to the patient are in the form of one or more dosage units, for example a tablet may be a single dosage unit and the container of a compound of Formula 1 in an aerosol may be a plurality of dosage units.

The substances used in the preparation of the pharmaceutical composition should be pharmaceutically pure and non-toxic in the amounts used. One of ordinary skill in the art will recognize that the optimal dosage of the active ingredient (s) in the pharmaceutical formulation depends on many factors. The factors include, but are not limited to, the type of subject (e.g., human), particular form of active ingredient, route of administration and formulation used. .

In general, the pharmaceutical composition comprises (where an indefinite article is used herein and throughout the specification, one or more times), one active ingredient according to Formula 1 as described herein, in admixture with one or more carriers. The carrier (s) may be in particulate form such that the compositions are, for example, in the form of a tablet or a powder. The carrier (s) may be liquid (s) and the compositions are, for example, an oral syrup or injectable liquid. In addition, the carrier (s) may be gaseous (s) to obtain an aerosol mixture useful, for example, by inhalation.

The composition intended for oral administration is preferably in solid or liquid form, and semi-solid, semi-liquid suspension and gel forms are included herein in forms contemplated as either solid or liquid.

The solid preparation for oral administration may be in the form of a powder, granule, compressed tablet, pill, capsule, chewing gum, wafer or the like. Such a solid preparation typically comprises one or more inert diluents or edible carriers. In addition, one or more of the following excipients may be used: binders such as carboxymethylcellulose, ethylcellulose, microcrystalline cellulose or gelatin, excipients such as starch, lactose or dextrins, disintegrants such as alginic acid, sodium alginate, Primogel, corn starch and similarly, lubricants such as magnesium stearate or Sterotex, glidants such as colloidal silica, sweeteners such as sucrose or saccharin, flavoring agents such as mint flavor, methyl salicylate or orange flavor, and coloring agents.

When the composition is in the form of a capsule, for example a gelatin capsule, it may contain, in addition to the substances of the type described above, a liquid carrier such as polyethylene glycol, cyclodextrin or a fatty oil.

The composition may be in the form of a liquid, for example an elixir, syrup, solution, emulsion or suspension. For example, the liquid may be for oral administration or for injection. When intended for oral administration, the preferred composition comprises, in addition to the compounds of the invention, one or more sweetening, preservative, coloring and flavoring agents. The formulation for injection may contain one or more surfactants, preservatives, wetting agents, dispersing agents, suspending agents, buffers, stabilizers, and isotonic agents.

The liquid pharmaceutical composition of the invention, when in the form of solutions, suspensions, or the like, may comprise one or more of the following excipients: sterile diluents such as water for injection, saline, preferably saline, Ringer's solution, isotonic sodium chloride solution, fixed oils such as synthetic mono or diglycerides which may serve as a solvent or suspending medium, polyethylene glycols, glycerol, cyclodextrin, propylene glycol or other solvents, antibacterial agents such as benzyl alcohol or methylparaben, antioxidants such as ascorbic acid or sodium bisulfite, chelating agents such as ethylenediaminetetraacetic acid, buffers such as acetates, citrates or phosphates and agents for adjusting the osmotic pressure such as sodium chloride or dextrose. The parenteral preparation can be enclosed in ampoules, disposable syringes or multi-dose vials made of glass or plastic. The preferred excipient is saline. The injectable pharmaceutical composition is preferably sterile. The liquid composition contemplated for parenteral or oral administration should contain an amount of a compound of Formula I that can be suitably dosed. Typically, this amount is at least 0.01% of the compound of the invention in the formulation. For an oral formulation, this amount may vary from 0.1% to about 80% by weight of the formulation. Preferred oral preparations contain between about 4% and about 50% active compound of formula 1. Preferred compositions and preparations of the invention are prepared in such a way that one parenteral dosage unit contains between 0.01% and 2% by weight of the active ingredient.

The pharmaceutical composition may be contemplated for topical administration, in which case the carrier may suitably include a solution, emulsion, ointment or gel base. For example, the base may include one of the following: natural petrolatum, lanolin, polyethylene glycols, beeswax, mineral oil, diluents such as water and alcohol, and emulsifiers and stabilizers. The pharmaceutical composition for topical administration may contain thickeners. For transdermal administration, the composition may include a transdermal patch or iontophoresis device. Conventional formulations may contain a concentration of a compound of Formula 1 of from about 0.1% to about 10% (weight per unit volume).

The composition may be contemplated for rectal administration, for example, in the form of a suppository that dissolves in the rectum to release the drug. The formulation for rectal administration may contain an oily base as a suitable non-irritating excipient. These bases include, but are not limited to, lanolin, cocoa butter, and polyethylene glycol.

The formulation may include a variety of agents that alter the physical form of the solid or liquid dosage unit. For example, the formulation may include substances that form a shell surrounding the active ingredients. The substances constituting this coating are usually inert and can be selected, for example, as sugar, shellac and other enteric coating agents. Alternatively, the active ingredients may be encapsulated in a gelatin capsule.

The formulation in solid or liquid form may include a formulation that binds the active ingredient (s) to assist in delivery of the active ingredients. Suitable agents that can function in this manner include monoclonal or polyclonal antibodies, proteins or liposomes.

The pharmaceutical composition of the invention may contain gaseous dosage units, for example, in the form of an aerosol. The term aerosol is used to refer to various systems ranging from those of a colloidal nature to systems representing pressurized packs. Delivery may be effected by liquefied or compressed gas or by a suitable pump system that delivers the active ingredients. Aerosols of the compounds of this invention may be delivered by a single-phase, two-phase, or three-phase system to ensure delivery of the active ingredient (s). The aerosol delivery device includes the necessary container, trigger device, valves, sub-containers, adapters, and the like, which may together form a kit. Preferred aerosols can be determined by one of ordinary skill in the art without undue experimentation.

The pharmaceutical composition of the invention, whether in solid, liquid or gaseous form, contains one or more pharmacological agents used in the treatment of inflammation (including asthma, allergy, rheumatoid arthritis, multiple sclerosis, etc.), proliferative disorders (carcinomas), diseases treatable by regulation calcium (including hypertension, cardiac arrhythmias, etc.) and acquired immune deficiency syndrome (AIDS).

The pharmaceutical compositions may be prepared by methods well known in the art of pharmacy.

A composition contemplated for injection may be obtained by mixing a compound of Formula 1 with water to form a solution. A surfactant may be added to facilitate the formation of a homogeneous solution or suspension. Surfactants are compounds that interact in a non-covalent manner with a compound of Formula 1 to facilitate the formation of a solution or homogeneous suspension of the active ingredient in an aqueous delivery system.

E. Biological effect

The compounds of formula I described herein or compositions comprising one or more of these compounds and a pharmaceutically acceptable carrier, diluent or excipient can be used in a method of treating or preventing an inflammatory condition or disease in a patient in which the method is: comprising administering to a patient in need of such treatment an amount of a compound or composition of the invention that is effective for treating or preventing an inflammatory condition or disease in the patient.

The inflammatory condition or disease may include respiratory inflammation (e.g., if the respiratory disease is asthma or if the respiratory disease is chronic obstructive pulmonary disease, or if the respiratory disease is emphysema), the inflammatory condition may be an autoimmune condition or disease, the inflammatory condition or disease may be lupus erythematous , the inflammatory condition or disease may include acute or chronic inflammation of the bone and / or cartilage joints of the joints, the inflammatory condition or disease may be arthritis selected from rheumatoid arthritis, gout arthritis or juvenile rheumatoid arthritis, the inflammatory condition or the disease may be the condition or disease may be accompanied by leukocyte infiltration, the condition or disease may be accompanied by swelling, the condition or disease may be accompanied by ischemic reperfusion injury, the condition or damaged ie, it may be accompanied by elevated levels of inflammatory cytokines (e.g., when the inflammatory cytokine is IL-4, or when the inflammatory cytokine is IL-5, or when the inflammatory cytokine is IL-10, or when the cytokine is IL-13, or when the cytokine is IL-9, or when the inflammatory cytokine is IL-1, or when the inflammatory cytokine is IL-2, or when the inflammatory cytokine is IL-6, or when the inflammatory cytokine is IL-18, or when is the inflammatory cytokine IL-3, or when the inflammatory cytokine is IL-8, or when the inflammatory cytokine is IL-12, or when the inflammatory cytokine is TNF-α, or when the inflammatory cytokine is TNF-β, or when the inflammatory cytokine is GM-CSF, or when the inflammatory cytokine is IFN-gamma, or when the inflammatory cytokine is LTB4, or when the inflammatory cytokine is a member of cysteinyl leuk otriene group, or when the inflammatory cytokine is regulated by activation of normal expressed and released T cells (RANTES), or when the anti-inflammatory cytokine is eotaxin-1,2 or 3, or when the inflammatory cytokine is a macrophage inflammatory protein (MIP) -1a, or when the inflammatory cytokine is a monocyte chemoattractive protein-1,2,3 or 4), the condition or disease may be accompanied by altered levels of inflammatory adhesive molecules (e.g., when the adhesive molecule is a vascular cell adhesive molecule (VCAM-1 or 2) when the adhesive molecule is an intracellular adhesive molecule (ICAM-1 or 2) when the adhesive molecule is a very late antigen-4 (VL_A-4) when the adhesive molecule is a very late antigen-1 related to leukocyte function when it is an adhesive a selectin molecule), the inflammatory condition in the disease may be multiple sclerosis, the inflammatory condition or the disease may be pulmonary sarcoidosis, the inflammatory condition or the disease may be eye inflammation or allergy, the inflammatory condition or the disease may be allergic rhinitis, the inflammatory condition or the disease may be an inflammatory bowel disease (e.g. Crohn's disease or ulcerative colitis), the inflammatory condition or the disease be an inflammatory skin disease (e.g., psoriasis or dermatitis), the inflammatory condition or disease may be graft versus host disease, the inflammatory condition or disease may be a vascular disease (e.g. vasculitis), the inflammatory condition or disease may be atherosclerotic disease.

Further, the present invention provides a method of treating or preventing a disease or condition in a patient in which the disease or condition is associated with a pathological condition that includes leukocyte infiltration, the method comprising administering to a patient in need thereof treatment an amount of a compound or is effective for treating or preventing a disease or condition associated with pathological conditions involving leukocyte infiltration.

Further, the present invention provides a method of treating or preventing asthma in a patient, comprising administering to said patient in need of such treatment an amount of a compound or composition of the invention that is effective for treating or preventing asthma in said patient.

Further, the present invention provides a method of treating or preventing allergy in a patient, comprising administering to said patient in need of such treatment an amount of a compound or composition of the invention that is effective for treating or preventing allergy in said patient.

In the method of the invention, a compound of Formula 1 or a composition comprising one or more compounds of Formula 1 and a pharmaceutically acceptable carrier, diluent or excipient may, but need not, achieve one or more of the following desired results in a patient being administered. a compound of formula 1 as defined above, or a composition comprising one of the following compounds and a pharmaceutically acceptable carrier, diluent or excipient:

1. inhibition of leukocyte infiltration (e.g. neutrophils, eosinophils, etc.),

2. inhibition of leukocyte activation

3. altering the ratio of lymphocytes (e.g. TH1 cells to TH2);

4. inhibition of leukocyte chemotaxis;

5. Inhibition of TNF-α production and / or release

6. inhibition of the formation and / or release of chemokines (eg eotaxin, etc.),

7. inhibiting the formation, release and / or function of an adhesive molecule (e.g., VCAM, VLA-4, etc.);

8. inhibition of edema;

9. inhibition of interleukin cytokine production and / or release (e.g. IL1, IL-2, IL-3, IL-4, IL-5, IL-6, IL-8, IL-9, IL-10, IL-12); IL-13, IL-18)

10. inhibiting the release of an inflammatory mediator (e.g., leukotrienes, tryptase, adenosine, etc.).

11. inhibition of histamine release;

12. suppression of asthma parameters a

13. suppression of allergy parameters.

The compounds of formula I described herein (i.e., compounds of formula 1 or compounds of this invention), or compositions comprising one or more of these compounds, a pharmaceutically acceptable carrier, diluent or excipient, can be used in a method of treatment or prevention. a proliferative disorder in a patient, comprising administering to the patient in need of such treatment an amount of a compound or composition of the invention that is effective to treat or prevent the proliferative disorder of the patient. The proliferative disorders contemplated herein include, without limitation, all leukemias and solid tumors that responsive to differentiation or apoptosis upon interruption of their cell cycle.

The compounds of formula I described herein (i.e., compounds of formula 1 or compounds of this invention), or compositions comprising one or more of these compounds, a pharmaceutically acceptable carrier, diluent or excipient, can be used in a method of treatment or prevention. diseases treatable by regulating calcium in a patient, the method comprising administering to a patient in need of such treatment an amount of a compound or composition of the invention effective to treat or prevent the disease in the patient. The term calcium-treatable disease includes, without limitation, cardiac arrhythmia, atrial fibrillation, acute coronary syndromes, hypertension, ischemic perfusion damage, stroke, epilepsy, demyelinating diseases such as multiple sclerosis, pain, epileptic condition, and atherosclerosis.

The compounds of formula I described herein (i.e., compounds of formula 1 or compounds of the present invention), or compositions comprising one or more of these compounds, a pharmaceutically acceptable carrier, diluent or excipient can be used in a method of treatment or prevention Acquired Immunodeficiency Syndrome (AIDS) in a patient, the method comprising administering to a patient in need of such treatment an amount of a compound or composition of the invention effective to treat or prevent the acquired immune deficiency syndrome in the patient. Acquired immune deficiency as used herein refers to infection by type 1 human immunodeficiency virus and includes, without limitation, associated complications such as acquired dementia complex syndrome and acquired immune deficiency syndromes.

Thus, the method of the invention can be used to treat inflammation, including acute and chronic inflammation, as well as certain proliferative disorders (cancer, diseases treatable by calcium regulation and AIDS). The term inflammation as used herein includes, without limitation, ankylosing spondylitis, arthritis (which includes 100 types of rheumatic diseases), asthma, chronic obstructive pulmonary disease, allergy, allergic rhinitis, Crohn's disease, fibromyalgia syndrome (gout, brain inflammation) including multiple sclerosis, AIDS dementia, Lyme encephalopathy, herpes encephalitis, Creutzfeld-Jakob disease and cerebral toxoplasmosis), emphysema, inflammatory bowel disease, irritable bowel syndrome, ischemic reperfusion injury, atopic dermatitis, pancreatitis, juvenile susceptible pseudosnuitis, juvenile susceptible pseudosusitis pelvic inflammatory disease, psoriatic arthritis (psoriasis), rheumatoid arthritis, psoriasis, tissue / organ transplantation, graft versus host disease, scleroderma, spondyloarthropathy, systemic lupus erythematosus, pulmonary sarcoidosis, mycopulitis, vasculitis, vasculitis, vasculitis, arditis and ulcerative colitis.

The method of the invention provides administration of a therapeutically effective amount of a compound of Formula I including salts, compositions, etc. thereof. The actual amount encompassed herein by the term "therapeutically effective amount" depends on the route of administration, the type of warm-blooded animal being treated and the physical characteristics of the animal under consideration. These factors and their relationship to the determination of this amount are well known to one of ordinary skill in the medical arts. This amount and route of administration can be adjusted to achieve optimal efficacy, but it depends on factors such as body weight, diet, concomitant use of other drugs, and other factors recognized by one of ordinary skill in the medical arts.

An effective amount of a compound or composition of the invention is sufficient to treat inflammation, proliferative disease, diseases treatable by calcium regulation or AIDS in a warm-blooded animal such as man. Methods of administering effective amounts of anti-inflammatory agents are well known in the art and include administration of inhaled, oral and parenteral forms. Such dosage forms include, but are not limited to, parenteral solutions, tablets, capsules, sustained release implantable forms and transdermal delivery systems, or inhalation delivery systems using dry powder inhalers or a multi-dose pressurized inhaler device.

The dosage amount and frequency are selected to produce an effective level without deleterious effects. In general, it will range from about 0.001 to 100 mg / kg / day and usually from 0.01 to 10 mg / kg / day for oral intravenous administration. Dosage ranges will also generally be from 0.0001 to 10 mg / kg / day for intranasal or inhalation administration.

Compounds of formula 1, including those used in the methods and compositions as defined above, can be prepared according to the schemes provided in the following examples. The following examples are offered by way of illustration and not by way of limitation.

DETAILED DESCRIPTION OF THE INVENTION

Unless otherwise stated, flash chromatography and column chromatography can be performed on Merck 60 silica gel. Flash chromatography can be performed according to the method described in Purification of Laboratory Chemicals, 3rd edition, Butterworth-Heinemann Ltd., Oxford (1988), red. DD Perrin and WLF Armatego, p. 23. Column chromatography refers to a method wherein the flow of eluent through the cartridge is by weight. Alternatively, flash chromatography and radial chromatography may alternatively be used in all cases. Radial chromatography is used on Chromatotron Model # 7924T silica gel (Harrison Reserch, Palo Alto, California). Unless otherwise stated, the values obtained are described _Rf TLC silica gel 60 F ₂ 54 (Merck KGaA, 64271, Darmstadt, Germany). Brine means saturated sodium chloride solution.

Chemical reactants and reagents are obtained, unless otherwise indicated, from standard chemical sources of supply such as Aldrich (Milwaukee, WI, www.aldrich.sial.com), EM Industries, Inc. (Hawthorne, NY, www.emscience.com), Fisher Scientific Co. (Hampton, NH, www.fischer1.com) and Lancaster Synthesis, Inc. (Windham, NH, www.lancaster.co.uk). Sulfo-NHS-biotin comes from Pierce (Rockford, IL, www.piercenet.com). MP-TsOH resin, PS-DIEA resin, PStrisamine resin, and PS-benzaldehyde resin are from Argonaut Technologies (San Carlos, CA, www.agrotech.com). The gases come from Praxair (Vancouver, B. C.). Cell lines, unless otherwise noted, come from public and commercial sources, for example, the American Tissue Culture Collection (ATCC, Rockville, MD).

Examples of preparation methods

Example 1

3-Amino-6,7-dihydroxy-17-ethylidene steroid

Compound 49, a representative compound of the invention, is prepared according to Scheme 1. Using a similar method, any number of compounds related to compound 49 can be obtained. Starting material, compound 45, can be prepared according to the method described in US patent 6,046,185. -bromide and potassium tert-butanol in toluene. Treatment of the 3β-hydroxyl compound 46 with ZnNe-2py, triphenylphosphine and DIAD in toluene affords the 3α-azide compound 47. Reduction of the azide with lithium aluminum hydride in diethyl ether affords the amine 48. Treatment with hydrochloric acid in tetrahydrofuran and water removes the acetonide group to form the ammonium 49.

Scheme 1

i) CH 3 CH ₂ PPh 3 Br, .KO'Bu, toluene, ii) ZnN6-2py, Ph ₃ P, DIAD, toluene, iii) LiAlH 4, Et ₂ O, iv) HCl, THF, water.

Preparation of Compound 46

A solution of tert-butanol potassium salt (0.24 g, 2.0 mmol), ethyltriphenylphosphine bromide (0.75 g, 2.0 mmol) and toluene (2.5 mL) was stirred at room temperature under argon. After 1 h, the dark red solution was cooled on ice and ketone (184 mg, 0.508 mmol) was added and the resulting solution was allowed to warm to room temperature. After stirring overnight, the reaction was quenched by the addition of 10 mL of water, diluted with 60 mL of ethyl acetate, the layers were separated and the organic layer was washed with 2 x 10 mL of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated. Purification by column chromatography eluting with 1: 1 ethyl acetate / hexane gave 171 mg (90%) of 46 as a colorless film.

Preparation of Compound 47

DIAD (0.44 mL, 2.14 mmol) was added dropwise over 10 min to a solution of 3β-hydroxyl compound 46 (400 mg, 1.07 mmol), ZnN ₆ -2py (246 mg, 0.80 mmol) , triphenylphosphine (560 mg, 2.14 mmol) and toluene (10.7 mL) at room temperature under an argon atmosphere. After 4 h, the reaction mixture was loaded onto a silica gel column with 10% ethyl acetate / hexane and eluted with 20% ethyl acetate / hexane to give 422 mg (99%) of compound 47 as a white solid.

Preparation of Compound 48

Lithium aluminum hydride (42 mg, 1.04 mmol) was added to an ice-cold solution of azide 47 (415 mg, 1.04 mmol) in 5.2 mL diethyl ether under argon. The reaction mixture was allowed to warm to room temperature. After 2 h, the solution was cooled in ice, diluted with 25 mL of diethyl ether, and slowly quenched by the addition of 2 mL of saturated sodium sulfate solution. After 10 min, a white precipitate formed and the solution was diluted with 50 mL of ethyl acetate, washed with 3 x 10 mL of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated. The crude material was purified on a silica gel column treated with 1% triethylamine / dichloromethane and washed with 5% methanol / dichloromethane. The crude material was added to the column in dichloromethane, eluting with 5% methanol / dichloromethane and then with dichloromethane: methanol: triethylamine 95: 5: 2 to give a white foam that contained triethylamine traces according to ¹ H NMR. The material was taken up in 50 mL of hexane, washed with 2 x 20 mL of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated to give 322 mg (83%) of 48 as a white solid foam.

Preparation of compound 49

A solution of the 3α-amino compound 48 (317 mg, 0.850 mmol), a 4 M solution of hydrochloric acid in dioxane (255 µ mmol, 1.02 mmol), tetrahydrofuran (13.6 mL) and water (3.4 mL) was stirred overnight at room temperature. The solution is evaporated to dryness, triturated with 3 x 10 ml acetone, after each trituration the acetone is evaporated. Evaporation gave 301 mg (96%) of 49 as a white solid.

LC / MS (direct injection, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 334.16, C ₂₁ H ₃₆ NO 2.

Example 2

3-Amino-6,7-dihydroxy-17-methylidene steroid

Another example of alkenes related to compound 49 is shown in Scheme 2. Olefination of ketone 45 using methyltriphenylphosphonium bromide and potassium tert-butanolate in tetrahydrofuran affords the 17-methylidene compound 50. Azidation with ZnN ₆ -2py, triphenylphosphine and DIAD in toluene provides the 3α-azide compound. 51. Reduction with lithium aluminum hydride in tetrahydrofuran affords the 3α-amino compound

52. Treatment with 80% acetic acid removes the acetonide protecting group and forms the ammonium acetate salt 53. Alternatively, compound 52 is treated with hydrochloric acid in acetonitrile and water to give the hydrochloride 54.

Scheme 2 o

HO ’ii

OH

i) CH ₃ PPh ₃ Br, KO'Bu, THF, ii) ZnN ₆ 2py, Ph ₃ P, DIAD, toluene, iii) LiAlH ₄ , Et ₂ O, iv) 80% acetic acid or water, MeCN.

Preparation of Compound 50

A solution of tert-butanol potassium salt (2.0 g, 16.9 mmol), methyltriphenylphosphonium bromide (6.0 g, 16.8 mmol) and TJ mL of tetrahydrofuran was stirred at room temperature under argon. After 1 h, ketone 45 (2.00 g, 5.52 mmol) was added to the yellow solution and the resulting solution was refluxed for 1 h. The reaction was quenched by the addition of 50 ml of saturated sodium chloride solution, diluted with 100 ml of ethyl acetate, the organic layer was separated and washed with 20 ml of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated. Purification by column chromatography eluting with ethyl acetate / hexane 1: 1 gave 1.90 g (95%) of Compound 50 as a white solid.

Preparation of Compound 51

DIAD (0.85 mL, 4.10 mmol) is added dropwise over 15 min at room temperature to a solution of 3β-hydroxyl compound 50 (739 mg, 2.05 mmol), ZnN6-2py (473 mg, 1.54) mmol), triphenylphosphine (1.075 g, 4.10 mmol) and toluene (20 mL) under an argon atmosphere. After 4 h, the reaction mixture was loaded onto a silica gel column packed with 10% ethyl acetate / hexane and eluted with 20% ethyl acetate / hexane to give 743 mg (94%) of compound 51 as a white foamy solid.

Preparation of Compound 52

Lithium aluminum hydride (77 mg, 1.93 mmol) was added to an ice-cooled solution of 3α-azide 51 in 5 mL of tetrahydrofuran and 5 mL of diethyl ether under an argon atmosphere. The reaction mixture was allowed to warm to room temperature. After 4 h, the solution was cooled in ice, diluted with 25 mL of diethyl ether and quenched slowly by the addition of 5 mL of saturated sodium sulfate solution. After 10 min, a white precipitate formed and the solution was diluted with 50 mL of ethyl acetate, washed with 3 x 10 mL of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated. The crude material was purified on silica gel packed with 1% triethylamine / dichloromethane and washed with 5% methanol / dichloromethane. The crude material was added in dichloromethane, eluting with 5% methanol / dichloromethane followed by dichloromethane: methanoctriethylamine 95: 5: 2 to give 636 mg (92%) of compound 52 as a white solid.

Preparation of Compound 53

A solution of 3α-amine 52 (287 mg, 0.799 mmol) and 10 mL of 80% acetic acid was heated at 40 ° C for 1 h. The reaction mixture was evaporated to give a white foam. Acetone (10 mL) was added and the solution was sonicated to dissolve and then evaporated. An additional 10 mL of acetone was added, the solution was sonicated and evaporated to give 301 mg (99%) of Compound 53 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water: acetonitrile) 320.19, C 20 H 34 NO 2.

Preparation of Compound 54

Add a solution of hydrochloric acid in dioxane to a solution of amine 52 in 1 ml of acetonitrile and 50 μΙ of water. The resulting rubbery solid was diluted with 2 mL of acetonitrile and vigorously stirred until a solid formed. The solid was filtered off and dried to give 50 mg (63%) of compound 54.

LC / MS mass spectrometry (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water: acetonitrile) 320.19 C20H34NO2.

Example 3

3-Amino-6,7-dihydroxy-17-methylidene steroid (alternative preparation)

Intermediate 52 is also prepared in an alternate manner as shown in Scheme 3. Azidation of alcohol 45 ZnN ₆ -2py, PPh ₃ and DIAD in toluene affords the 3aazide compound 55. Hydrogenation of azide in the presence of palladium on carbon provides amine 56. Olefination of compound 56 with methyltriphenylphosphonium bromide and potassium the tert-butanol salt in tetrahydrofuran affords the 17-methylidene compound 52.

Scheme 3

i) ZnN _with -2py, Ph ₃ P, DIAD, toluene, ii) H ₂ , Pd, EtOAc, iii) CH ₃ PPh ₃ Br, KO'Bu, THF.

Preparation of Compound 55

DIAD (2.4 mL, 11.6 mmol) was added dropwise over 20 min to a solution of 3β-hydroxyl compound 45 (2.108 g, 5.81 mmol), ZnN ₆ -2py (1.34 g, 4.36) mmol), triphenylphosphine (3.05 g, 11.6 mmol) and toluene (58 mL) at room temperature under an argon atmosphere. The reaction was allowed to proceed overnight and the mixture was poured onto a silica gel column and eluted with 20% ethyl acetate / hexane to give 1.14 g (50%) of compound 55 as a white foamy solid.

Preparation of Compound 56

A solution of azide 55 (1.10 g, 2.84 mmol), 10% palladium on carbon (60 mg, 0.057 mmol) and 28 mL of ethyl acetate was stirred at room temperature overnight under a hydrogen atmosphere. The solution was filtered through celite and eluted with ethyl acetate. Purification by radial chromatography eluting with dichloromethane: methanol: triethylamine 95: 5: 2 gave 892 mg (81%) of compound 56 as a white solid.

Preparation of Compound 52

A solution of tert-butanol potassium salt (175 mg, 1.48 mmol), methyltriphenylphosphonium bromide (528 mg, 1.48 mmol) and 3 mL of tetrahydrofuran was stirred at room temperature under argon. After 1 h, ketone 56 (100 mg, 0.277 mmol) was added to the yellow solution and the resulting solution was allowed to stir at room temperature overnight. The reaction is quenched by the addition of 5 ml of water, diluted with 50 ml of ethyl acetate, the aqueous layer is separated and washed with 10 ml of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated. Purification by radial chromatography eluting with dichloromethane: methanol: triethylamine 95: 5: 2 gave 96 mg (97%) of compound 52 as a white solid.

Example 4

3-Amino-6,7-dihydroxy-17-fluoroethylidene steroid

Halogen analogs similar to compound 49 can be prepared from halogenated olefinic compositions. Scheme 4 shows the preparation of the 20-fluorinated analog 64. The hydroxyl group of compound 45 is protected by treatment with tert-butyldimethylsilyl chloride and imidazole in dimethylformamide. Olefination of ketone 57 using triethyl (2-fluoro-2-phosphonoacetate) anion provides a mixture of compound 58 and its geometric isomer. The compounds can be separated by silica gel chromatography. Reduction of the ester with lithium aluminum hydride in diethyl ether affords allyl alcohol 59. A mixture of sulfur trioxide pyridine complex in tetrahydrofuran followed by addition of lithium aluminum hydride affords the dehydroxylated compound 60. in toluene affords the 3α-azide compound 62. Reduction with lithium aluminum hydride in tetrahydrofuran gives 3α-amine 63. Hydrochloric acid treatment in tetrahydrofuran and water removes the protecting groups from the hydroxyls at the 6- and 7- positions to give the ammonium chloride salt.

Scheme 4

El

iv) SO ₃ .Py, THF, LiAlH ₄ , v) Bu ₄ NF, THF, vi) ZnN ₆ -2py, Ph ₃ P, DIAD, toluene, vii) LiAlH ₄ , Et ₂ O, viii) HCl, THF, Water.

Preparation of Compound 57

A solution of ketone 45 (4.73 g, 13.1 mmol), TBSCI (3.01 g, 19.6 mmol), imidazole (2.67 g, 39.2 mmol) and dimethylformamide (52 mL) was stirred at temperature room overnight. The white suspension was diluted with 250 mL of ethyl acetate, washed with 2 x 50 mL of water and 50 mL of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated to give 5.97 g (96%) of compound 57 as a white solid.

Preparation of Compound 58

Lithium bis (trimethylsilyl) amide (10.0 mL of a 1.0 M solution in tetrahydrofuran) (10.0 mL) was added to a solution of (EtO) ₂ P (O) -CHFCO ₂ Et (2.65 g, 10, 5 mmol) at room temperature (2.65 g, 10.5 mmol) in tetrahydrofuran (22 mL) under argon. After 1 h, a solution of ketone 57 (2.50 g, 5.25 mmol) in tetrahydrofuran (20 mL) was added and the resulting solution was heated at reflux for 4.5 h and then stirred at room temperature overnight. The reaction was quenched by the addition of 1.5 mL of saturated sodium bicarbonate solution and then partially evaporated to remove most of the tetrahydrofuran. The residue is diluted with 200 ml of ethyl acetate, washed with 3 x 20 ml of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated. The crude material was purified by column chromatography eluting

2.5% then 5% ethyl acetate / hexane to give 1.50 g (50%) of compound 58 as a white solid.

Preparation of Compound 59

Lithium aluminum hydride (106 mg, 2.66 mmol) was added to an ice-cold solution of ester 58 (1.50 g, 2.66 mmol) in diethyl ether (13 mL) under an argon atmosphere. The solution was allowed to warm to room temperature. After 3 h, the solution is cooled in ice and 20 ml of saturated sodium sulfate solution are added slowly. After 10 min, the solution was diluted with 150 mL of ethyl acetate, washed with water and saturated brine, dried over magnesium sulfate, filtered and evaporated to give 1.44 g (quantitative yield) of compound 59 as a white foam.

Preparation of Compound 60

The sulfur trioxide pyridine complex (69.5 mg, 0.428 mmol) was added to an ice-cooled solution of allyl alcohol 59 (149 mg, 0.285 mmol) in tetrahydrofuran (2.8 mL) under an argon atmosphere. After 6 h, lithium aluminum hydride (68 mg, 1.71 mmol) was added and the solution was allowed to warm to room temperature overnight. The solution is cooled in ice and 5 ml of saturated sodium sulfate solution are slowly added. After 10 min, the solution was diluted with 75 mL of ethyl acetate, washed with water and saturated brine, dried over magnesium sulfate, filtered and evaporated to give 109 mg (76%) of compound 60 as a white solid.

Preparation of Compound 61

A solution of compound 60 (410 mg, 0.810 mmol), tetrabutylammonium fluoride (0.89 mL of a 1.0 M solution in tetrahydrofuran, 0.89 mmol), and tetrahydrofuran (5 mL) was refluxed under argon. After 1.5 h, the solution was cooled to room temperature, diluted with 75 mL of ethyl acetate, washed with 20 mL of water and 2 x 20 mL of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated.

The residue was filtered through silica gel eluting with ethyl acetate and evaporated to give 318 mg (100%) of compound 61 as a white solid.

Preparation of Compound 62

DIAD (0.33 mL, 1.59 mmol) was added dropwise over 10 min to a solution of 3β-alcohol 61 (312 mg, 0.796 mmol), ZnN ₆ -2py (183 mg, 0.597 mmol), triphenylphosphine (417 mg) , 1.59 mmol) and toluene (8.0 mL) at room temperature under an argon atmosphere. After 3 h, the reaction mixture was transferred to a silica gel column packed with 10% ethyl acetate / hexane and eluted with 20% ethyl acetate / hexane to give 322 mg (97%) of compound 62 as a crystalline solid.

Preparation of Compound 63

Lithium aluminum hydride (29 mg, 0.75 mmol) was added to an ice-cooled solution of azide 62 (314 mg, 0.753 mmol) in 7.5 mL of diethyl ether under argon. The reaction mixture was allowed to warm to room temperature and stirred overnight. The solution was cooled in ice and the reaction was slowly quenched by the addition of 10 mL of saturated sodium sulfate solution. After 10 min, a white precipitate formed and the solution was diluted with 75 mL of ethyl acetate, washed with 20 mL of water and 2 x 20 mL of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated. The crude material was purified on a silica gel column treated with 1% triethylamine / dichloromethane and washed with 5% methanol / dichloromethane. The crude material was taken up in dichloromethane, eluting with 5% methanol / dichloromethane followed by dichloromethane / methanol / triethylamine 95: 5: 2 to give a white solid. ¹ H NMR showed traces of triethylamine and therefore the material was taken up in 75 mL of dichloromethane and washed with 2 x 25 mL of water, dried over magnesium sulfate, filtered and evaporated to give 137 mg (47%) of 63 as a colorless film .

Preparation of Compound 64

A solution of 3α-amino compound 63 (137 mg, 0.35 mmol), 4 M hydrochloric acid in dioxane (105 μΙ, 0.42 mmol), tetrahydrofuran (5.6 mL) and water (1.4 mL) was stirred at room overnight. The solution was evaporated, the residue was taken up twice with 3 ml of methanol and evaporated to give 130 mg (96%) of compound 64 as an off-white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 352.14, C21H35FNO2.

Example 5

3-Amino-6,7-dihydroxy-17-carbomethoxyethylidene steroid

Olefination of compounds similar to compound 57 can also be performed to provide 21-carboalkoxy-substituted analogs. Scheme 5 shows the preparation of a 21-carbomethoxy-substituted compound 69 as an example. Olefination of ketone 57 using trimethyl (2-phosphonoacetate) anion provides a mixture of compound 65 and its geometric isomer. Compounds can be separated by silica gel chromatography. Tetrabutylammonium fluoride in tetrahydrofuran removes the protecting group from the 3-hydroxyl group to give compound 66. Azidation using ZnN ₆ -2py, triphenylphosphine and DIAD in toluene affords 3α-azido compound 67. Hydrogenation of azide in the presence of palladium on carbon as catalyst provides 3α-amine. Treatment with 80% acetic acid removes hydroxyl protecting groups at the 6 and 7 positions to give the ammonium acetate salt 69.

(i) (MeO) ₂ P (O) CHFCO ₂ Me, LiN (TMS) ₂ , THF, (ii) Bu ₄ NF, THF, (iii) ZnN ₆ -2py, Ph ₃ P,

DIAD, toluene, iv) H ₂ Pd, EtOAc, v) 80% acetic acid

Preparation of Compound 65

Lithium bis (trimethylsilyl) amide (2.00 mL of a 1.0 M solution in tetrahydrofuran, 2.00 mmol) was added to a solution of (MeO) ₂ P (O) CH ₂ Me (390 mg, 2.10 mmol) in tetrahydrofuran (22 mL) at room temperature under argon. After 3 h, a solution of ketone 57 (509 mg, 1.07 mmol) in tetrahydrofuran (2 mL) was added and the resulting solution was refluxed for 3 days. The reaction was quenched by the addition of 5 mL of water, diluted with 75 mL of ethyl acetate, washed with 2 x 15 mL of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated. The crude material was purified by column chromatography eluting with 5% ethyl acetate / hexane to afford 307 mg (54%) of compound 65 as a colorless film. 119 mg (21%) of the Z isomer is also isolated.

Preparation of Compound 66

A solution of compound 65 (296 mg, 0.550 mmol), tetrabutylammonium fluoride (0.61 mg of a 1.0 M solution in tetrahydrofuran, 0.61 mmol), and tetrahydrofuran (3 mL) was refluxed under argon. After 1 h, the solution was cooled to room temperature, diluted with 20 mL of ethyl acetate, washed with 10 mL of water and 2 x 10 mL of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated to afford 230 mg (100%) of compound 66. in the form of a white solid.

Preparation of Compound 67

DIAD (252 µL, 1.28 mmol) was added dropwise to a solution of 3β-alcohol 66 (230 mg, 0.55 mmol), ZnN ₆ -2py (147 mg, 0.48 mmol), triphenylphosphine (335 mg, 1 mmol) , 28 mmol) and toluene (6.4 mL) at room temperature under argon. After 2 h, the reaction mixture was purified by radial chromatography eluting with 15% ethyl acetate / hexane to afford 203 mg (84%) of compound 67.

Preparation of Compound 68

A solution of azide 67 (203 mg, 0.45 mmol), 10% palladium on carbon (48 mg, 0.045 mmol), and 4.5 mL of ethyl acetate was stirred at room temperature under a hydrogen atmosphere for 3 days. The solution was filtered through celite and eluted with ethyl acetate and methanol to afford 165 mg (88%) of Compound 68.

Preparation of Compound 69

A solution of amine 68 (165 mg, 0.40 mmol) and 2 mL of 80% acetic acid was heated at 40 ° C for 1 h. The reaction mixture was diluted with 10 mL of toluene and then evaporated to remove the residual acetic acid. Trituration of the residue in 10 mL of cyclohexane followed by filtration and drying afforded 117 mg (67%) of compound 69 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in water / acetonitrile 4: 1) 378.17, C22H36NO4.

Example 6

3α-Amino-6α, 7-dihydroxyandrostatin-17-one acetate salt

In an analogous manner, compounds having different functional groups on a C17 carbon atom can be obtained. For example, a 17-ketone compound is obtained by treating compound 56 with 80% acetic acid to give the 3α-amino-6α, 7β-dihydroxyandrostan-17-one acetate salt 70 (see Table 2).

Preparation of Compound 70

A solution of ketone 56 (67 mg, 0.16 mmol) and 1 mL of 80% acetic acid was heated at 40 ° C for 1 h. The reaction mixture was diluted with 10 mL of toluene and then evaporated to give 63 mg (100%) of compound 70.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in water / acetonitrile 4: 1) 322.18, C ₁₉ H 32 NO ₃ .

Example 7

3-Amino-6,7-dihydroxy-17-hydroxy steroid

Analogs substituted with the hydroxy group at position 17 can be prepared from ketones similar to compound 56 as shown in Scheme 6. The carbonyl group in compound 56 is reduced with sodium borohydride in methanol to give exclusively the 17β-hydroxy isomer 71. Treatment with 80% acetic acid removes the acetonide protecting group to form the ammonium salt. 72nd

Scheme 6

i) NaBH ₄ , MeOH; ii) 80% acetic acid

Preparation of Compound 71

The ice-cold solution of ketone 56 (100 mg, 0.28 mmol), sodium borohydride (16 mg, 0.41 mmol) and 1.4 mL of methanol was allowed to react for 2.5 h. The reaction was quenched by the addition of 1 mL of water and the mixture was evaporated to remove most of the methanol. The residue was diluted with 40 mL of dichloromethane and washed with 2 x 10 mL of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated to give 90 mg (90%) of compound 71.

Preparation of Compound 72

A solution of amine 71 (90 mg, 0.25 mmol) and 2 mL of 80% acetic acid was heated at 40 ° C for 2 h. The reaction mixture was diluted twice with 10 mL of toluene and evaporated to remove the residual acetic acid. The remaining material was dissolved in 1 mL of methanol and 5 mL of hexane, evaporated and dried to give 86 mg (90%) of compound 72 as a white solid.

LC / MS mass spectrometry (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 324.19, C19H34NO3.

Example 8

Salts of 3α-amino-6,7-dihydroxy-17-methylidenensteroid

The hydroxyl groups at the 6 and 7 positions can be protected by various protecting groups. Suitable protecting groups are listed in the Greene and Wuts Protective Groups in Organic Synthesis, John Wiley & Sons, New York, N.Y. (1999). Scheme 7 shows examples of analogous compounds that can be prepared with hydroxyl groups at the 6 and 7 positions protected as methyl ethers. The starting compound 73 is disclosed in U.S. Pat. The formation of the dianion of compound 73 using sodium hydride in dimethylformamide followed by alkylation with methyl iodide affords compound 74. Treatment with 80% acetic acid removes cyclic ketal and tert-butyldimethylsilyl ether protecting groups. Olefination of compound 75 with methyltriphenylphosphonium bromide and potassium tert-butanol in tetrahydrofuran affords the 17-methylidene compound 76. Azidation with ZnN6-2py, triphenylphosphine and DIAD in toluene affords the 3α-azide compound 77. hydrochloric acid in diethyl ether and methanol affords the ammonium chloride salt 79. Treatment of compound 78 with acetic acid leads to the ammonium acetate salt 80.

Scheme 7

i) NaH, Mel, DMF, ii) 80% acetic acid, iii) CH ₃ PPh ₃ Br, KO ¹ Bu, THF, iv) ZnNg-2py, Ph ₃ P, DIAD, toluene, v) LiAlH ₄ , Et ₂ 0, vi) HCl, Et ₂ O, MeOH or acetic acid

Preparation of Compound 74

Sodium hydride (0.50 g, 12.4 mmol) was added to a solution of room temperature diol 73 (1.49 g, 3.10 mmol) in 10 mL of dimethylformamide under a nitrogen atmosphere. After 2 h, the solution was cooled in ice and methyl iodide (1.93 mg, 30.9 mmol) was added dropwise over 30 s. The reaction mixture was allowed to warm to room temperature with stirring overnight. The reaction mixture was diluted with 100 mL of diethyl ether, washed with 10 mL of water and 2 x 10 mL of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated to give 1.73 g of crude compound 74 as a pale yellow oil.

Preparation of Compound 75

A solution of crude compound 74 (1.73 g, 3.10 mmol) and 15 mL of 80% acetic acid was stirred at room temperature for 4 h. The solution is evaporated, the residue is taken up in 50 ml of ethyl acetate, washed with 2 x 20 ml of saturated sodium bicarbonate solution and 10 ml of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated to give 1.18 g of crude compound 75 as white foamy substance.

Preparation of Compound 76

A solution of tert-butanol potassium salt (1.09 g, 9.20 mmol), methyltriphenylphosphonium bromide (3.30 g, 9.20 mmol) and 15 mL of tetrahydrofuran was stirred at room temperature under a nitrogen atmosphere. After 2 h, crude ketone 75 (1.17 mg, 3.08 mmol) was added to the yellow solution and the resulting solution was allowed to stir at room temperature overnight. The reaction was quenched by the addition of 2 ml of water, diluted with 100 ml of ethyl acetate, washed with 3 x 10 ml of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated. Column chromatography eluting with 80% ethyl acetate / hexane gave 89 mg of impure compound 76 as a white solid.

Preparation of Compound 77

DIAD (1.05 mg, 5.08 mmol) is added dropwise over 10 min to a solution of 3β-alcohol 76 (885 mg, 2.54 mmol), ZnN6-2py (585 mg, 1.90 mmol), triphenylphosphine (1.33 g, 5.08 mmol) and toluene (25 mL) under an argon atmosphere. After 11 h, the reaction mixture was purified by column chromatography eluting with 15% ethyl acetate / hexane to afford 594 mg (63%) of compound 77 as a crystalline solid.

Preparation of Compound 78

Lithium aluminum hydride (0.79 mL of a 1 M solution in diethyl ether, 0.79 mmol) was added to an ice-cold solution of azide 77 (588 mg, 1.57 mmol) in 15.7 mL of diethyl ether under an argon atmosphere. After 10 min, the reaction mixture was allowed to warm to room temperature with stirring overnight. After 1 h, the reaction mixture was cooled in ice, and the reaction was slowly quenched by the addition of 10 mL of saturated sodium sulfate solution. After 10 min, a white precipitate formed and the liquid was decanted. The residue was washed with 2 x 25 mL of ethyl acetate and the washings were combined with the previously decanted ether solution. The solution was washed with saturated sodium chloride solution (3 x 10 mL), dried (MgSO 4), filtered and evaporated. Purification by column chromatography eluting with dichloromethane / methanol / triethylamine 95: 5: 2 afforded 434 mg (79%) of compound 78 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 348.20, C22H38NO2.

Preparation of compound

Hydrochloric acid (0.26 mL of a 1.0 M solution in diethyl ether, 0.26 mmol) was added to a solution of amine 78 (60 mg, 0.17 mmol) in 2 mL of diethyl ether. The resulting gel-like substance was dissolved in 5 ml of methanol and evaporated. The residue was dissolved in 1 mL of methanol, diluted with 5 mL of cyclohexane and evaporated to give 66 mg (100%) of compound 79 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 348.20, C22H38NO2.

Preparation of Compound 80

A solution of amine 78 (61 mg, 0.17 mmol) and 1 mL of acetic acid was allowed to stand at room temperature for 30 min. The solution is diluted with 5 ml of toluene and evaporated. The residue was taken up in 5 mL of hexane, evaporated and the residue was dried for 2 h in an Abderhalden refluxing acetone dryer to give 71 mg (100%) of compound 80 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 348.20, C22H38NO2.

Example 9

Salts of 3β-amino-6,7-dihydroxy-17-methylidenensteroid

The stereochemistry at C3 can be converted by adding the 3β-ammonium salt derivative of any compound similar to compound 49. The stereochemistry at C3 can be converted in three steps, as shown in Scheme 8 for the preparation of compounds 28 and 83. The 3β-hydroxyl compound 46 is converted to ββ-mesylate 81 with methanesulfonyl chloride and pyridine. Heating compound 81 and cesium acetate in dimethylformamide at 100 ° C affords the 3-acetate compound 82. The transformation sequence is terminated by methanolysis of the acetate of compound 82 with sodium methoxide to give the 3α-hydroxyl compound 25. Treatment of 25 ZnN ₆ -2py, triphenylphosphine and DIAD in toluene. reduction of the azide with lithium aluminum hydride in diethyl ether affords 3β-3ΐτιΐηοζΙύόβηϊηυ 27. Treatment with hydrochloric acid in tetrahydrofuran and water removes the acetonide group to give the ammonium chloride salt 28. Similarly, the acetone salt treatment of compound 27 yields 80% acetic acid. using the methods depicted in Scheme 80, compound 50 is converted to compound 89 and compound 61 is converted to compound 95 (see Table 1). Compounds 26, 27, 87, 88, 89, 93, 94, and 95 are examples of compounds of this invention with the stereochemistry of 3β.

Scheme 8

i) MsCl, pyridine, ii) CsOAc, DMF, 100 ° C, iii) NaOMe, MeOH, iv) ZnN ₆ -2py, Ph ₃ P, DIAD, toluene, v) LiAlH ₄ , Et ₂ O, vi) 4 M HCl in dioxane, THF, water or 80% acetic acid.

Preparation of Compound 81

Methanesulfonyl chloride (1.2 mL, 16 mmol) was added to an ice-cold solution of 3β-hydroxyl compound 46 (3.0 g, 8.0 mmol) in pyridine (20 mL) under argon. After 4 h, the solution is cooled in ice and 20 ml of saturated sodium bicarbonate solution are added. After 15 min, the solution was diluted with 150 mL of ethyl acetate and washed with 3 x 25 mL of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated to give 3.6 g (100%) of compound 81 as an off-white foam.

Preparation of Compound 82

A solution of mesylate 81 (3.6 g, 8.0 mmol), cesium acetate (4.6 g, 24 mmol) and 40 mL of dimethylformamide was heated at 100 ° C for 24 h. The solution was diluted with 100 mL of water, extracted with 2 x 100 mL of diethyl ether, washed with 2 x 50 mL of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated to give about 3 g of crude compound 82.

Preparation of Compound 25

A solution of sodium (398 mg, 17.3 mmol) in methanol (21.5 mL) was added to 3α-acetate 82 (1.8 g, 4.3 mmol) in tetrahydrofuran (10 mL). After 2 h, 20 mL of water was added and the resulting solution was diluted with 100 mL of ethyl acetate, washed successively with saturated sodium bicarbonate solution, water and saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated to give 1.58 g (98%). of crude compound 25 as a yellow foam.

Preparation of Compound 26

DIAD (1.70 mL, 8.24 mmol) was added dropwise over 10 min to a solution of 3α-alcohol 25 (1.54 g, 4.12 mmol), ZnNs-2py (0.94 g, 3.09) mmol), triphenylphosphine (2.16 g, 8.24 mmol) and toluene (44 mL) under an argon atmosphere. After stirring overnight, the reaction mixture was poured onto a silica gel column packed with 10% ethyl acetate / hexane and eluted with 10% ethyl acetate / hexane to give 0.89 g (61%) of 26 as a white solid.

Preparation of compound 27

Lithium aluminum hydride (146 mg, 3.66 mmol) was added to an ice-cooled solution of azide 26 (1.46 g, 3.66 mmol) in 18.3 mL diethyl ether under an argon atmosphere. The reaction mixture was allowed to warm to room temperature. After 1.5 h, the solution was cooled in ice, diluted with 25 mL of diethyl ether and quenched slowly by the addition of 20 mL of saturated ammonium sulfate solution. After 10 min, a white precipitate formed and the solution was diluted with 50 mL of ethyl acetate, washed with 3 x 10 mL of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated to give 1.31 g (96%) of compound 27 as white foamy substance.

Preparation of compound 28

A solution of 3β-amino compound 27 (227 mg, 0.609 mmol), a 4 M solution of hydrochloric acid in dioxane (183 μΙ, 0.73 mmol), tetrahydrofuran (9.7 mL) and water (2.4 mL) was stirred at room temperature. during the night. The tetrahydrofuran and water were evaporated, the residue was taken up in 5 ml of methanol and evaporated, triturated with 5 ml of acetone and evaporated to give 224 mg (100%) of compound 28 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 334.10, C21H36NO2.

Preparation of Compound 83

A solution of amine 27 (412 mg, 1.10 mmol) and 5 mL of 80% acetic acid was stirred at room temperature for 4 h. The reaction mixture is diluted with 5 ml of toluene and then evaporated. The residue is taken up in 2 x 5 ml of toluene and evaporated to remove the remaining acetic acid. The residue was triturated twice with 10 mL of dichloromethane and evaporated to give 430 mg (99%) of compound 83 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 334.19, C 21 H 36 NO 2.

Preparation of Compound 84

Methanesulfonyl chloride (0.33 mL, 4.2 mmol) was added to an ice-cooled solution of 3β-hydroxyl compound 50 (754 mg, 2.09 mmol) in pyridine (5.3 mL) under an argon atmosphere. After 4 h, the solution is cooled in ice and 5 ml of saturated sodium bicarbonate solution are added. After 15 min, the solution was diluted with 60 mL of ethyl acetate and washed 3 times with saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated to give 860 mg (94%) of compound 84 as an off-white solid.

Preparation of Compound 85

A solution of mesylate 84 (860 mg, 1.96 mmol), cesium acetate (1.13 g, 5.88 mmol) and 10 mL of dimethylformamide was heated at 95 ° C for 32 h. The solution is diluted with 50 ml of water, extracted with 2 x 100 ml of diethyl ether, washed with 2 x 30 ml of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated. Purification by column chromatography eluting with 5% and 8% ethyl acetate / hexane gave 558 mg (71%) of compound 85 as a white solid.

Preparation of Compound 86

A solution of sodium (128 mg, 5.56 mmol) in methanol (7 mL) was added to 3α-acetate 85 (558 mg, 1.38 mmol). After 2 h, 5 mL of saturated sodium bicarbonate solution was added and the resulting solution was diluted with 100 mL of ethyl acetate. The solution was washed with 2 x 20 mL of water and 2 x 20 mL of saturated sodium chloride solution and dried over magnesium sulfate, filtered and evaporated to give 491 mg (99%) of compound 86 as a white solid.

Preparation of Compound 87

DIAD (0.57 mL, 2.74 mmol) was added dropwise over 15 min to a solution of 3α-hydroxyl compound 86 (493 mg, 1.37 mmol), ZnN ₆ -2py (315 mg, 1.03 mmol) , triphenylphosphine (718 mg, 2.74 mmol) and toluene (13.7 mL) at room temperature under an argon atmosphere. After 3.5 h, the reaction mixture was poured onto a silica gel column packed with 10% ethyl acetate / hexane and eluted with 10% ethyl acetate / hexane to give 390 mg (74%) of 87 as a viscous oil.

Preparation of compound 88

Lithium aluminum hydride (40 mg, 1.01 mmol) was added to an ice-cooled solution of azide compound 87 (390 mg, 1.01 mmol) in 5 mL of diethyl ether under argon. The reaction mixture was allowed to warm to room temperature. After 2 h, the solution was cooled in ice, diluted with 25 mL of diethyl ether and slowly quenched with 2 mL of saturated sodium sulfate solution. After 10 min, a white precipitate formed and the solution was diluted with 40 mL of ethyl acetate, washed with 3 x 15 mL of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated. The crude material was purified by column chromatography on silica gel treated with 1% triethylamine / dichloromethane and washed with 5% methanol / dichloromethane. The crude material was taken up in dichloromethane, eluting with 5% methanol / dichloromethane and then 95: 5: 2 dichloromethane / methanol / triethylamine to afford 277 mg (76%) of 88 as a white solid.

Preparation of compound 89

A solution of the amino compound 88 (270 mg, 0.752 mmol) and 10 mL of 80% acetic acid was heated at 40 ° C for 1 h. The reaction mixture was evaporated to give a white foamy solid. Acetone (10 ml) was added and the mixture was sonicated to dissolve the substances and then evaporated. An additional portion of 10 mL of acetone was added, the mixture was sonicated and evaporated to give 285 mg (100%) of compound 89 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 320.26, C20H34NO2.

Preparation of Compound 90

Methanesulfonyl chloride (0.20 mol, 2.56 mmol) was added to an ice-cooled solution of ββ-hydroxyl compound 61 (501 mg, 1.28 mmol) in pyridine (3.2 mL) under an argon atmosphere. After 4 h, the solution is cooled in ice and 5 ml of saturated sodium bicarbonate solution are added. After 15 min, the solution was diluted with 50 mL of ethyl acetate and washed 3 times with saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated to give 590 mg (98%) of compound 90 as a white foamy solid.

Preparation of Compound 91

A solution of mesylate 90 (590 mg, 1.25 mmol), cesium acetate (722 mg, 3.76 mmol) and 6.2 mL of dimethylformamide was heated at 100 ° C for 24 h. The solution was diluted with 50 ml of water, extracted with 2 x 50 ml of diethyl ether, washed with 2 x 30 ml of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated. Purification by column chromatography eluting with 8% ethyl acetate / hexane gave 297 mg (55%) of compound 91 as a white solid.

Preparation of compound 92

A solution of sodium (63 mg, 2.7 mmol) in methanol (3.4 mL) was added to 3α-acetate 91 (297 mg, 0.684 mmol) in tetrahydrofuran (1 mL). The solution was stirred overnight, 5 mL of water and 80 mL of ethyl acetate were added and the mixture was washed twice with water and twice with saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated to afford 251 mg (94%) of 92 as a white. solids.

Preparation of compound 93

DIAD (0.26 mL, 1.24 mmol) was added dropwise over 10 min to a solution of 3α-alcohol 92 (243 mg, 0.620 mmol), ZnN ₆ -2py (143 mg, 0.465 mmol), triphenylphosphine (325 mg). , 1.24 mmol) and toluene (6.2 mL) at room temperature under an argon atmosphere. After 4 h, the reaction mixture was loaded onto a silica gel column packed with 10% ethyl acetate / hexane and eluted with 20% ethyl acetate / hexane to give 209 mg of impure compound 93 as a yellow oil.

Preparation of Compound 94

Lithium aluminum hydride (20 mg, 0.50 mmol) was added to ice-cooled crude azide 93 (209 mg, 0.50 mmol) in 5 mL of diethyl ether under nitrogen. The reaction mixture was allowed to warm to room temperature. After 4 h, the solution was cooled in ice, diluted with 25 mL of diethyl ether and quenched slowly by the addition of 2 mL of saturated sodium sulfate solution. After 10 min, a white precipitate formed and the solution was diluted with 50 mL of ethyl acetate, washed with 3 x 10 mL of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated. The crude material is chromatographed using a silica gel column treated with 1% triethylamine / dichloromethane and washed with 5% methanol / dichloromethane. The crude material was loaded onto a dichloromethane column, eluting with 5% methanol / dichloromethane followed by 95: 5: 2 dichloromethane / methanol / triethylamine to give 97 mg of impure compound 94 as a white solid.

Preparation of compound 95

A solution of impure 3β-amino compound 94 (97 mg, 0.25 mmol), 4 M hydrochloric acid in dioxane (74 μΙ, 0.30 mmol), tetrahydrofuran (4 mL), and water (1 mL) was stirred at room temperature. After 4 h the solution was evaporated, the residue was taken up in 5 ml of methanol and evaporated. The residue was triturated twice with 5 ml of acetone and evaporated. Dissolve the white solid in about 0.5 mL of water and slowly add acetone (5 mL) until crystals appear. The crystals were filtered off, washed with acetone and dried to give 66 mg of compound 95 as colorless fine needles.

LC / MS mass spectrometry (direct infusion, electrospray + ve, 10 mM ammonium acetate solution in water / acetonitrile 4.9) 352.09, C21H35FNO2.

Example 10

3-Amino-6,7-dihydroxy-17-ALKYL

Any 17 (20) -alkenyl functional group can be hydrogenated on a double bond with hydrogen in the presence of a catalyst such as 10% palladium on carbon. For example, compound 96 of compound 28 can be prepared as shown in Scheme 9. Similarly, compound 97 of compound 49 can be prepared as shown in Scheme 9 (see Table 2).

Scheme 9

i) H ₂ , Pd on carbon, methanol.

Preparation of compound 96

A solution of olefin 28 (52 mg, 0.14 mmol), 10% palladium on carbon (15 mg, 0.014 mmol) and methanol (3 mL) was stirred overnight at room temperature under a hydrogen atmosphere. The solution was filtered through celite, eluting with 50 mL of methanol and evaporated to give 50 mg (96%) of compound 96 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 336.24, C 21 H 38 NO 2.

Preparation of Compound 97

A solution of olefin 49 (844 mg, 2.28 mmol), 10% palladium on carbon (243 mg, 0.228 mmol) and methanol (11 mL) was stirred at room temperature overnight under a hydrogen atmosphere. The solution was filtered through celite, eluting with 50 ml of methanol and evaporated. The residue was triturated with 10 mL of acetone, filtered and dried to give 801 mg (94%) of compound 97 as a white solid.

LC / MS mass spectrometry (direct infusion, electrospray + ve, 10 mM ammonium acetate solution in water / acetonitrile 4: 1) 336.21 C 21 H ₃₈ NO ₂ .

Example 11

3-sec-amino-6,7-dihydroxy-17-METHYLIDENE

Any amine similar to compound 52 can be coupled to an aldehyde or ketone to provide secondary or tertiary amines. Treatment of compound 52 with a solution of 4-isopropylbenzaldehyde and titanium isopropoxide in tetrahydrofuran followed by reduction with sodium borohydride affords compound 99. Treatment with 8% acetic acid removes the acetonide group to give the ammonium acetate salt 100. Examples of compounds 101-107 are prepared as shown in 10 Table 6).

Scheme 10

i) 4-isopropylbenzaldehyde, Ti (O'Pr) 4, THF, NaBH ₄ , MeOH, ii) 80% acetic acid

Preparation of Compound 99

Titanium isopropoxide (120 μΙ, 0.42 mmol) was added to a solution of amine 52 (100 mg, 0.28 mmol), 4-isopropylbenzaldehyde (46 μΙ, 0.31 mmol) and 1.4 mL of tetrahydrofuran at room temperature below. nitrogen atmosphere. After 12 h, a solution of sodium borohydride (29 mg, 0.78 mmol) in 1 mL of ethanol was added and the reaction was allowed to proceed for an additional 8 h. The reaction was quenched by the addition of 3 mL of saturated sodium chloride solution, diluted with 30 mL of ethyl acetate, partitioned, washed with 10 mL of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated. Purification by radial chromatography gave 50 mg (36%) of compound 99.

Preparation of Compound 100

A solution of amine 99 (50 mg, 0.10 mmol) and 1 mL of 80% acetic acid was heated at 40 ° C for 3 h. The reaction mixture was taken up with two 5 ml portions of toluene and evaporated and then once with acetone and hexane to give 25 mg (51%) of the compound.

100th

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 452.27, C30H46NO2.

Preparation of Compound 101

Following the procedure described for the preparation of compound 99, amine 52 (100 mg, 0.28 mmol) was reacted with 2-fluorobenzaldehyde (32 µL, 0.32 mmol) to give 43 mg of the amine intermediate. This amine intermediate was treated with 1 mL of 80% acetic acid at 40 ° C for 3 h. The reaction mixture was diluted with 5 mL of toluene and evaporated. The residue was dissolved in 1 mL acetone, diluted with 5 mL hexane and evaporated to give 49 mg (37%) of compound 101 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 428.22, C27H39FNO2.

Preparation of Compound 102

Following the procedure described for the preparation of compound 99, amine 52 (100 mg, 0.28 mmol) was reacted with 3- (trifluoromethyl) benzaldehyde (41 µL, 0.31 mmol) to give 61 mg of the amine intermediate. This amine intermediate was treated with 1 mL of 80% acetic acid at 40 ° C for 3 h. The reaction mixture was diluted with 5 mL of toluene and evaporated. The residue was dissolved in 1 mL of acetone, diluted with 5 mL of hexane and evaporated to give 64 mg (45%) of compound 102 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 478.18, C28H39F3NO2.

Preparation of Compound 103

Following the procedure described for the preparation of compound 99, amine 52 (100 mg, 0.28 mmol) was reacted with o-anisaldehyde (42 mg, 0.31 mmol) to give 30 mg of the amine intermediate. This amine intermediate was treated with 1 mL of 80% acetic acid at 40 ° C for 3 h. The reaction mixture was diluted with 1 mL of toluene and evaporated. The residue was dissolved in 1 mL of acetone, diluted with 5 mL of hexane and evaporated to give 18 mg (14%) of compound 103 as a white solid.

LC / MS mass spectrometry (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 440.23, C 28 H 42 NO 3.

Preparation of Compound 104

Following the procedure described for the preparation of compound 99, amine 52 (100 mg, 0.28 mmol) was reacted with 4- (trifluoromethoxy) benzaldehyde (44 μΙ, 0.31 mmol) to give 86 mg of the amine intermediate. This amine intermediate is treated with 1.5 mL of 80% acetic acid at 40 ° C for 3 h. The reaction mixture was diluted with 1 mL of toluene and evaporated. The residue was dissolved in 1 mL acetone, diluted with 5 mL hexane and evaporated to give 84 mg (57%) of compound 104 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 494.15, C28H39NO3.

Preparation of compound 105

Following the procedure described for the preparation of compound 99, amine 52 (100 mg, 0.28 mmol) was reacted with 3-phenoxybenzaldehyde (60 mg, 0.32 mmol) to give 73 mg of the amine intermediate. This amine intermediate was treated with 1 mL of 80% acetic acid at 40 ° C for 3 h. The reaction mixture was diluted with 5 mL of toluene and evaporated. The residue was dissolved in 1 mL of acetone, diluted with 5 mL of hexane and evaporated to give 87 mg (58%) of compound 105 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in water / acetonitrile 4: 1) 502.20, C 33 H 44 NO 3.

Preparation of compound 106

Following the procedure described for the preparation of compound 99, amine 52 (100 mg, 0.28 mmol) was reacted with 3-nitrobenzaldehyde (46 mg, 0.31 mmol) to give 18 mg of the amine intermediate. This amine intermediate was treated with 1 mL of 80% acetic acid at 40 ° C for 3 h. The reaction mixture was diluted with 5 mL of toluene and evaporated.

The residue was dissolved in 1 mL acetone, diluted with 5 mL hexane and evaporated to give 18 mg (14%) of compound 106 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 455.20, C27H39N2O4.

Preparation of Compound 107

Following the procedure described for the preparation of compound 99, amine 52 (200 mg, 0.55 mmol) was reacted with 3-pyridylcarboxaldehyde (82 µg, 0.61 mmol) to give 100 mg of the amine intermediate. A suspension of the amine intermediate, a 4 M solution of hydrochloric acid in dioxane (65 µL, 0.26 mmol), 110 µL of water, and 2.2 mL of acetonitrile was stirred at room temperature for 1 h. The solution was filtered and the solid dried to give 77 mg (30%) of compound 107 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate solution in water / acetonitrile 4: 1) 411.21, C 26 H 39 N 2 O 2.

Example 12

3-cycloamino-6,7-dihydro-17-etylidénsteroid

Any ketone similar to compound 108 can be coupled to an amine as shown in Scheme 11. Starting material, compound 108 for preparation is described in U.S. Patent 6,046,185. Reaction of compound 108 with piperidine and sodium cyanoborohydride in methanol affords compound 109 as a mixture of isomers on C3. Treatment with 80% acetic acid removes the acetonide protecting groups and forms the ammonium acetate salt 110. An example of compound 111 is prepared by the methods shown in Scheme 11 except that hydrochloric acid is used in place of acetic acid (see Table 5). A 3-cycloamino group is a group attached at the 3-position wherein the carbon at the 3-position is directly attached to the nitrogen and that nitrogen is part of the heterocyclic ring.

Scheme 11 ο

? H

110 en 7- '7- Ο / 10 »I

(i) piperidine, Ti (O'Pr) 4, THF, NaBH ₄ , MeOH); (ii) 80% acetic acid

Preparation of compound 109

A solution of ketone 108 (200 mg, 0.54 mmol), piperidine (266 μΙ, 2.68 mmol), 100 mg of 0.3 nm molecular sieves, sodium cyanoborohydride (24 mg, 0.38 mmol) and 5.4 mL of methanol was added. Stir at room temperature for 24 h. The reaction mixture was diluted with 20 mL of water and extracted with 2 x 20 mL of dichloromethane. The combined extracts were washed with 10 ml of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated. The crude material was purified by radial chromatography eluting with 20% methanol / dichloromethane to afford 112 mg (47%) of compound 109 as a white solid.

Preparation of Compound 110

A solution of amines 109 (102 mg, 0.23 mmol) and 5 mL of 80% acetic acid was heated at 40 ° C for 1 h. The solution is evaporated, the residue is taken up in 2 ml of methanol, diluted with 15 ml of toluene and evaporated. The residue was triturated with 5 mL of acetone, filtered and dried to give 44 mg (42%) of compound 10 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 402.31, C 26 H 44 NO ₂ .

Preparation of Compound 111

In the manner described for the preparation of compound 109, ketone 108 (200 mg

0.54 mmol) with morpholine (234 μΙ, 2.68 mmol) to give 56 mg of the amine intermediate. This amine intermediate was treated with 5 mL of 80% acetic acid at 40 ° C for 1 h. The solution was evaporated, dissolved in 5 ml of methanol and evaporated again. ¹ H and ¹³ C NMR indicated removal of the acetonide protecting group, but little or no salt was formed. This material was treated with a 4 M solution of hydrochloric acid in dioxane (32 μΙ, 0.13 mmol) and 2 mL of acetone to give a white precipitate. The suspension was diluted with 2 mL of acetone, filtered and dried to give 48 mg (20%) of compound 111 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 404.20, C 25 H 42 NO 3.

Example 13

Conversion of the 3-oxo group in the steroid to the 3-sec-amino group

Any ketone similar to compound 108 can be coupled to an amine as shown in Scheme 12. Methylamine is added to a solution of compound 108 and titanium isopropoxide in tetrahydrofuran, followed by reduction with sodium borohydride. The solution is filtered and eluted on MP-TsOH resin to give compound 112 as a mixture of isomers on C3. Treatment with hydrochloric acid in acetonitrile and water affords the ammonium chloride salt 113. Examples of compounds 114 to 129 are prepared by the methods depicted in Scheme 12, except that acetic acid is used in place of hydrochloric acid in cases where ammonium acetate salts are formed (cf. Table 5).

i) methylamine hydrochloride, Ti (O'Pr) 4, THF, NaBH ₄ , MeOH, ii) HCl, water, acetonitrile.

Preparation of Compound 112

Titanium isopropoxide (270 µL, 0.92 mmol) was added to a solution of room temperature ketone 108 (250 mg, 0.67 mmol), methylamine hydrochloride (41 mg, 0.61 mmol) and 1.5 mL of tetrahydrofuran under atmosphere. nitrogen. After 12 h, a solution of sodium borohydride (65 mg, 1.7 mmol) in 2.3 mL of ethanol was added and the reaction continued for another 10 h. The reaction was quenched by the addition of 0.5 mL of water and the mixture was filtered to remove a white precipitate. The solution was applied to a column of 600 mg of MP-TsOH resin and eluted with 3 mL of methanol followed by 4 mL of a 2M solution of ammonia in methanol. The ammonia / methanol reaction was evaporated to give 76 mg (32%) of compound 112.

Preparation of Compound 113

A suspension of compound 112 (76 mg, 0.21 mmol), 4 M hydrochloric acid in dioxane (75 μΙ, 0.30 mmol), 50 μΙ water and 1 mL acetonitrile was stirred at room temperature for 1 h. The solution was filtered and the solid dried to give 38 mg (13%) of Compound 113 as a gray solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in water / acetonitrile 4: 1) 348.19, C ₂₅ H ₃₈ NO ₂ .

Preparation of Compound 114

Using the methods described for the preparation of compound 112, ketone 108 (200 mg, 0.53 mmol) is reacted with propylamine hydrochloride (47 mg, 0.49 mmol) to give 72 mg of the amine intermediate. This amine intermediate was treated with 75 μΙ of hydrochloric acid solution to give 34 mg (17%) of Compound 114.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 376.22, C2 ₄ H ₄₂ NO ₂ .

Preparation of Compound 115

Using the methods described for the preparation of compound 112, ketone 108 (250 mg, 0.67 mmol) is reacted with amylamine (70 µΙ, 0.61 mmol) to give 82 mg of the amine intermediate. This amine intermediate was treated with 75 µl of hydrochloric acid solution to give 75 mg (28%) of compound 115 as a white solid.

LC / MS mass spectrometry (direct infusion, electrospray + ve, 10 mM ammonium acetate solution in water / acetonitrile 4: 1) 404.28, C ₆ H _{4 6} NO ₂ .

Preparation of Compound 116

Using the methods described for the preparation of compound 112, ketone 108 (250 mg, 0.67 mmol) is reacted with cyclopentylamine (60 µΙ, 0.61 mmol) to give 99 mg of the amine intermediate. This amine intermediate is treated with 200 μΙ of acetic acid for 1 h and taken up twice with 1 ml of toluene and evaporated. The residue was triturated with 1 mL of cyclohexane, filtered and dried to give 98 mg (35%) of compound 116.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 402.27, C26H44NO2.

Preparation of Compound 117

Using the methods described for the preparation of compound 112, ketone 108 (250 mg, 0.67 mmol) is reacted with cyclohexylamine (70 µΙ, 0.61 mmol) to give 120 mg of the amine intermediate. This amine intermediate was treated with 0.5 mL of acetic acid for 1 h and taken up twice with 1 mL of toluene and evaporated. The residue was triturated with 1 mL of cyclohexane, filtered and dried to give 101 mg (32%) of compound 117.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate solution in water / acetonitrile 4: 1) 416.25, C27H46NO2.

Preparation of Compound 118

Using the methods described for the preparation of compound 112, ketone 108 (250 mg, 0.67 mmol) was reacted with pyrrolidine (51 µL, 0.61 mmol) to give 70 mg of the amine intermediate. This amine intermediate was treated with 50 µl of acetic acid for 1 h and 1 mL of cyclohexane was added to give a solid which was filtered and dried to give 65 mg (22%) of 118.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 388.28, C 25 H 42 NO 2.

Preparation of Compound 119

Using the methods described for the preparation of compound 112, ketone 108 (200 mg, 0.53 mmol) was reacted with N-propylethylenediamine (60 µL, 0.49 mmol) to give 99 mg of the amine intermediate. This amine intermediate was treated with 75 µl of acetic acid to give 47 mg (20%) of compound 119.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 419.32, C 26 H 47 NO 2.

Preparation of Compound 120

Using the methods described for the preparation of compound 112, ketone 108 (250 mg, 0.67 mmol) was reacted with N, N-dimethylethylenediamine (65 µL, 0.61 mmol) to give 93 mg of the amine intermediate. This amine intermediate was treated with 75 µl hydrochloric acid solution over 1 h to afford 77 mg (29%) of compound 120 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 405.28, C25H45NO2.

Preparation of Compound 121

Using the procedures described for the preparation of compound 112, ketone 108 (250 mg, 0.67 mmol) was reacted with piperazine (52 mg, 0.61 mmol) to give 33 mg of the amine intermediate. This amine intermediate was treated with 200 µl acetic acid for 1 h and 1 µl of cyclohexane was added and twice taken and evaporated with 1 ml of toluene. The residue was triturated with 1 mL of cyclohexane, filtered and evaporated to give 39 mg (14%) of compound 121.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 403.23, C 25 H 43 N 2 O 2.

Preparation of Compound 122

Using the methods described for the preparation of compound 112, ketone 108 (250 mg, 0.67 mmol) was reacted with ethanolamine (33 µΙ, 0.61 mmol) to give 136 mg of the amine intermediate. This amine intermediate was treated with 75 μΙ of hydrochloric acid solution to give 124 mg (50%) of Compound 122 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 378.19, C23H40NO3.

Preparation of Compound 123

Using the procedures described for the preparation of compound 112, ketone 108 (250 mg, 0.67 mmol) was reacted with 5-amino-1-pentanol (63 mg, 0.61 mmol) to give 129 mg of the amine intermediate. This amine intermediate was treated with 50 μΙ of hydrochloric acid solution to give 65 mg (24%) of Compound 123 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 420.25, C26H46NO3.

Preparation of Compound 124

Using the methods described for the preparation of compound 112, ketone 108 (250 mg, 0.67 mmol) was reacted with 2- (2-aminoethylamino) ethanol (62 μΙ, 0.61 mmol) to give 90 mg of the amine intermediate. This amine intermediate was treated with 75 μΙ of hydrochloric acid solution to give 79 mg (28%) of Compound 124 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 421.24, C25H45N2O3.

Preparation of Compound 125

Using the methods described for the preparation of compound 112, ketone 108 (200 mg, 0.53 mmol) was reacted with m-toluidine (52 µL, 0.49 mmol) to give 95 mg of the amine intermediate. This amine intermediate is treated with 75 μΙ of hydrochloric acid solution to give 43 mg (19%) of Compound 125.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 424.23, C28H42NO2.

Preparation of Compound 126

Using the methods described for the preparation of compound 112, ketone 108 (250 mg, 0.67 mmol) was reacted with 4-aminophenol (67 mg, 0.61 mmol) to give 138 mg of the amine intermediate. This amine intermediate was treated with 75 μΙ of hydrochloric acid solution to give 41 mg (14%) of 126.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 426.18, C27H40NO3.

Preparation of Compound 127

Using the methods described for the preparation of Compound 112, ketone 108 (250 mg, 0.67 mmol) was reacted with sulfanilamide (105 mg, 0.61 mmol) to afford, after purification by radial chromatography, 24 mg of the amine intermediate. This amine intermediate was treated with 75 μΙ of hydrochloric acid solution to give 23 mg (7%) of Compound 127.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in water / acetonitrile 4: 1) 489.17, C 27 H ₄₁ N 2 O ₄ S.

Preparation of Compound 128

Using the methods described for the preparation of compound 112, ketone 108 (250 mg, 0.67 mmol) was reacted with 3-aminomethylpyridine (62 μΙ, 0.61 mmol) to give 108 mg of the amine intermediate. This amine intermediate was treated with 1 mL of 80% acetic acid at 40 ° C for 1 h. The reaction mixture was evaporated and taken up twice with 1 ml portions of toluene and evaporated. The residue was triturated with 1 mL of cyclohexane, filtered and dried to give 117 mg (41%) of compound 128.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 425.24, C27H41N2O2.

Preparation of Compound 129

Using the methods described for the preparation of compound 112, ketone 108 (250 mg, 0.67 mmol) was reacted with histamine (68 mg, 0.61 mmol) to give 120 mg of the amine intermediate. This amine intermediate was treated with 1 mL of 80% acetic acid at 40 ° C for 1 h. The reaction mixture was evaporated and taken up twice with 1 mg portions of toluene and evaporated. The residue was triturated with 1 mL of cyclohexane, filtered and dried to give 128 mg (38%) of compound 129.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 428.23, C26H42N3O2.

Example 14

Conversion of the 3-amino group of the steroid to the 3-acylamino group

The amide and sulfonamide analogs can be prepared from any amine similar to compound 52. Scheme 13 shows the preparation of amide 131. Acetylation of amine 52 in dichloromethane using acetyl chloride and diethylamine bound to resin provides amide 130. Treatment with 80% acetic acid removes the acetonide group to afford dihydroxyamide 131.

Scheme 13 o

131

(i) acetyl chloride, PS-DIEA, CH2Cl2, PS-trisamine, (ii) 80% acetic acid

Preparation of Compound 130

A solution of amine 52 (100 mg, 0.28 mmol), acetyl chloride (50 µl, 0.70 mmol), 440 mg of PS-DIEA resin and 2.4 mL of dichloromethane was stirred at room temperature for 16 h. The resin is filtered and the filtrate is incubated for 2 h with 260 mg of PS-trisamine resin. The resin was filtered off and the filtrate was evaporated. Purification by radial chromatography afforded 69 mg (62%) of Compound 130.

Preparation of Compound 131

A solution of amide 130 (69 mg, 0.17 mmol) and 1 mL of 80% acetic acid was heated at 40 ° C for 1 h. The reaction mixture is taken up twice with 5 ml of toluene, evaporated, taken up once with 5 ml of methanol and evaporated and once with 1 ml of acetone and 5 ml of hexane and evaporated to give 62 mg (62%) of Compound 131 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in water / acetonitrile 4: 1) 384.16, C 22 H ₃₅ NNaO3, 362.20, C22H36NO3, 344.18 C22H34NO2.

Preparation of Compound 132

Following the procedure described for the preparation of compound 130, amine 52 (88 mg, 0.24 mmol) was reacted with benzoyl chloride (65 µL, 0.56 mmol) to give 64 mg of the amide intermediate. A solution of the amide intermediate and 2 mL of 80% acetic acid was heated at 40 ° C for 1 h. The reaction mixture is taken up twice with 5 ml of toluene and evaporated, once with 5 ml of methanol and once with 1 ml of acetone and 5 ml of hexane and evaporated to give 55 mg (55%) of compound 132 as a white solid (see Table 3).

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in water / acetonitrile 4: 1) 446.18, C 27 H _{3 3} 7NNaO, 424.29, C27H38NO3, 406.19, C27H36NO2.

Preparation of Compound 133

Following the procedure described for the preparation of compound 130, amine 52 (100 mg, 0.28 mmol) was reacted with isopropylsulfonyl chloride (63 μΙ, 0.56 mmol) to give 38 mg of the amide intermediate. A solution of the amide intermediate and 1.5 mL of 80% acetic acid was heated at 40 ° C for 1 h. The reaction mixture is taken up twice with 5 ml of toluene and evaporated, once with 5 ml of methanol and once with 1 ml of acetone and 5 ml of hexane and evaporated to give 35 mg (29%) of compound 133 as an off-white solid (see Table 3).

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 426.14, C23H40NO4S.

Preparation of Compound 134

Following the procedure described for the preparation of compound 130, amine 52 (100 mg, 0.28 mmol) was reacted with benzenesulfonyl chloride (90 μΙ, 0.70 mmol) to give 105 mg of the amide intermediate. A solution of the amide intermediate and 2 mL of 80% acetic acid was heated at 40 ° C for 1 h. The reaction mixture is taken up twice with 5 ml of toluene and evaporated, once with 5 ml of methanol and evaporated and once with 1 ml of acetone and 5 ml of hexane and evaporated to give 83 mg (65%) of Compound 134 as a white solid (see Table 3). )

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in water / acetonitrile 4: 1) 482.11, _C2 6H3 ₇ NNaO4S, 477.17, C ₂₆ H4iN ₂ O4S, 460.15 , C26H38NO4S.

Example 15

3-Acylaminobiotín-6,7-hydroxy-17-etylidénsteroid

Scheme 14 shows the preparation of amide 135. Reaction of amine 83 with triethylamine and a type of water-soluble N-hydroxysuccinimide biotinester in methanol and water provides a biotinylated amide analog 135.

Scheme 14

i) Sulfo-NHS-biotin, 13 N, MeOH, water.

Preparation of Compound 135

A solution of compound 83 (97 mg, 0.25 mmol), triethylamine (104 µΙ, 0.75 mmol), sulfo-NHS-biotin (120 mg, 0.27 mmol), 2.5 mL methanol and 2.5 mL water is stirred at room temperature overnight. The reaction mixture was evaporated and purified by reverse phase column chromatography eluting with 5% water / methanol to give 89 mg (64%) of Compound 135 as an off-white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 560.30, C31H50N3O4S.

Example 16

3-urea-6,7-hydroxy-17-METHYLIDENE

Any of the amines similar to compound 52 can be reacted with isocyanates or isothiocyanates to provide compounds having urea or thiourea functional groups. Compounds 136, 137 and 138 are examples of ureas prepared by the methods depicted in Scheme 15 (see Table 3).

Scheme 15

137

i) phenyl isocyanate, PS-Trisamine, CH ₂ Cl ₂ , ii) 80% acetic acid

Preparation of Compound 136

A solution of amine 52 (100 mg, 0.28 mmol), phenyl isocyanate (76 μΙ, 0.70 mmol) and 2.4 mL of dichloromethane was stirred at room temperature for 16 h. The solution is incubated for 2 h with 260 mg of PS-Trisamine resin. The resin was filtered off and the filtrate was evaporated. Purification by radial chromatography gave 95 mg (71%) of compound 136.

Preparation of Compound 137

A solution of urea 136 (95 mg, 0.20 mmol) and 2 mL of 80% acetic acid was heated at 80 ° C for 2 h. The reaction mixture is taken up in 5 ml of toluene and evaporated, 5 ml of methanol and evaporated and 5 ml of hexane and evaporated. Purification by radial chromatography eluting with dichloromethane / methanol / triethylamine 95: 5: 2 gave 40 mg (33%) of compound 137.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in water / acetonitrile 4: 1) 461.18, C ₂₇ H3eN ₂ NaO _3, 439.22, C ₂₇ H 9 N ₃ O _{2 3} , 421.25, C ₂₇ H ₃₇ N ₂ O ₂ .

Preparation of Compound 138

Following the procedure described for the preparation of compound 137, amine 52 (100 mg, 0.28 mmol) was reacted with propyl isocyanate (52 μΙ, 0.56 mmol) to give 72 mg of the urea intermediate. A solution of the urea intermediate and 2 mL of 80% acetic acid was heated at 80 ° C for 2 h. The reaction mixture was taken up twice with 5 ml of toluene and evaporated, once with 5 ml of methanol and evaporated and once with 5 ml of hexane and evaporated to give 51 mg (45%) of compound 138 as a white solid.

LC / MS mass spectrometry (direct infusion, electrospray + ve, 10 mM ammonium acetate solution in water / acetonitrile 4: 1) 427.21, C ₂₄ H ₄₀ N ₂ NaO 3, 405.25, C ₂₄ H ₄ iN ₂ O ₃ .

Example 17

Unsaturated 3-amino-6,7-hydroxy-17-dimethylsteroid

Any compounds related to compounds 88 or 89 can be rearranged using the method depicted in Scheme 16. Treatment of compound 88 with a solution of hydrochloric acid in methanol and water at 50 ° C removes the acetonide protecting groups, allows the 18-methyl group to migrate to the C17 position and form the ammonium chloride. Treatment of compound 89 under the same conditions also provides compound 139. The compounds of Examples 140-148 were prepared by the methods shown in Scheme 16 (see Table 4).

Scheme 16

i) HCl, water, methanol, 50 ° C

Preparation of Compound 139

A solution of compound 88 (300 mg, 0.834 mmol), 4 drops of concentrated hydrochloric acid, 2 mL of methanol and 2 mL of water was heated at 50 ° C for 72 h. The reaction mixture is evaporated and the residue is taken up twice with 5 ml of methanol and evaporated. The remaining material is taken up in 2 ml of methanol, diluted with 15 ml of acetone and evaporated. The residue was triturated with 5 mL of acetone, filtered and dried to give 286 mg (96%) of Compound 139 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 320.20, C20H34NO2.

Preparation of Compound 139

In the same manner as described for the preparation of compound 139 from compound 88, compound 89 was reacted to afford 145 mg (77%) of compound 139 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 320.20, C20H34NO2.

Preparation of Compound 140

Following the procedure described for the preparation of compound 99, amine 88 (200 mg, 0.55 mmol) was reacted with m-tolualdehyde (90 µL, 0.61 mmol). Purification by radial chromatography with 5% methanol / ethyl acetate gave 127 mg of the amine intermediate. A solution of the amine intermediate, 4 drops of concentrated hydrochloric acid, 1 mL of methanol and 1 mL of water was heated at 50 ° C for 20 h. The reaction mixture was taken up three times with 5 ml of methanol and evaporated and once with 5 ml of acetone and evaporated to give 74 mg (30%) of compound 140 as an off-white foam.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate solution in water / acetonitrile 4: 1) 424.24, C 28 H 42 NO 2.

Preparation of Compound 141

As described for the preparation of compound 99, amine 88 (200 mg, 0.55 mmol) was reacted with 3,4-difluorobenzaldehyde (67 μΙ, 0.61 mmol). Purification by radial chromatography with 30% ethyl acetate / hexane gave 88 mg of the amine intermediate. A solution of the amine intermediate, 4 drops of concentrated hydrochloric acid, 1 mL of methanol and 1 mL of water was heated at 50 ° C for 20 h. The reaction mixture was taken up three times with 5 ml of methanol and evaporated and once with 5 ml of acetone and evaporated to give 73 mg (28%) of compound 141 as an off-white foam.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 446.42, C27H38F2NO2.

Preparation of Compound 142

As described for the preparation of compound 99, amine 88 (200 mg, 0.55 mmol) was reacted with 3,4-dimethoxybenzaldehyde (70 μΙ, 0.61 mmol). Purification by radial chromatography with 40% ethyl acetate / hexane gave 67 mg of the amine intermediate. A solution of the amine intermediate, 4 drops of concentrated hydrochloric acid, 1 mL of methanol and 1 mL of water was heated at 50 ° C for 20 h. The reaction mixture was taken up three times with 5 ml of methanol and evaporated and once with 5 ml of acetone and evaporated to give 43 mg (16%) of compound 142 as a yellow solid.

LC / MS mass spectrometry (direct infusion, electrospray + ve, 10 mM ammonium acetate solution in water / acetonitrile 4: 1), 470.27, C 29 H 44 NO 4.

Preparation of Compound 143

A solution of compound 28 (200 mg, 0.540 mmol), 4 drops of concentrated hydrochloric acid and 3 mL of water was heated at 50 ° C for 72 h. The reaction mixture is evaporated and the residue is taken up twice with 5 ml of methanol and evaporated. The residue is taken up in 3 ml of methanol, diluted with 20 ml of acetone and evaporated. The remaining material was triturated with 10 mL of acetone, filtered and evaporated to give 179 mg (90%) of Compound 143 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 334.20, C 21 H 36 NO 2.

Preparation of Compound 144

Titanium isopropoxide (270 μΙ, 0.92 mmol) was added to a solution of ketone 108 (250 mg, 0.67 mmol), aniline (56 μΙ, 0.61 mmol) and 1.5 mL of tetrahydrofuran at room temperature under an argon atmosphere. . After 12 h, a solution of sodium borohydride (65 mg, 1.7 mmol) in 2.3 mL of ethanol was added and the reaction continued for an additional 8 h. The reaction was quenched by the addition of 0.5 mL of water and the mixture was filtered to remove a white precipitate. The solution was applied to a column of 600 mg of MP-TsOH resin and eluted with 9 mL of methanol and then 9 mL of a 2 M solution of ammonia in methanol. The ammonia / methanol fraction was evaporated and the residue was taken up in 4 ml of tetrahydrofuran and treated with 500 mg of PS-benzaldehyde resin and filtered to remove any residual aniline. Evaporate the solution and remove the residue with 2 ml of a 9: 1 mixture of tetrahydrofuran / water and 100 μΙ of concentrated hydrochloric acid. After stirring at room temperature overnight, the reaction mixture was evaporated.

The residue was triturated with 1 mL of cyclohexane, filtered and dried to give 62 mg of compound 144 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 410.03, C27H40NO2.

Preparation of Compound 145

Following the procedure described for the preparation of compound 144, compound 108 (250 mg, 0.67 mmol) was reacted with 3- (trifluoromethyl) aniline. The intermediate is purified by radial chromatography and then treated with 100 µl of concentrated hydrochloric acid in 2 ml of tetrahydrofuran / water 9: 1. After stirring at room temperature overnight, the reaction mixture was evaporated and the residue was triturated with 1 mL of cyclohexane, filtered and dried to give 23 mg of 145 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 477.94, C28H39F3NO2.

Preparation of Compound 146

Titanium isopropoxide (216 μΙ, 0.73 mmol) was added to a solution of ketone 108 (200 mg, 0.54 mmol), benzylamine (53 μΙ, 0.49 mmol) and 1.2 mL of tetrahydrofuran at room temperature under an argon atmosphere. . After 12 h, a solution of sodium borohydride (52 mg, 1.4 mmol) in 1.7 mL of ethanol was added and the reaction continued for an additional 6 h. The reaction was quenched by the addition of 1 mL of water and the mixture was filtered to remove a white precipitate. The solution is diluted with 70 ml of dichloromethane, washed with 10 ml of water and 20 ml of saturated sodium chloride solution, dried over magnesium sulfate, filtered and evaporated. Purification by radial chromatography, eluting successively with 20% ethyl acetate / hexane, ethyl acetate, and dichloromethane / methanol / triethylamine 95: 5: 2, gave 127 mg of the 3α-amine intermediate and 26 mg of the 3β-amine intermediate. A solution of 127 mg of the 3α-amine intermediate, 1 mL of tetrahydrofuran / water 9: 1 and 0.1 mL of concentrated hydrochloric acid was stirred overnight at room temperature. The reaction mixture is evaporated, the residue is taken up in 5 ml of methanol and evaporated. The remaining material was triturated with 1 mL of cyclohexane, filtered and dried to give 118 mg (95%) of Compound 146 as a white solid.

LC / MS mass spectrometry (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 424.20 C28H42NO2.

Preparation of Compound 147

A solution of 26 mg of the 3β-amine intermediate described above for the preparation of compound 146, 1 mL of 9: 1 hydrofuran / water and 0.1 mL of concentrated hydrochloric acid was stirred at room temperature overnight. The reaction mixture is evaporated, the residue is taken up in 5 ml of methanol and evaporated. The residue was triturated with 1 mL of cyclohexane, filtered and dried to give 26 mg (100%) of Compound 147 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 424.21, C 28 H 42 NO 2.

Preparation of Compound 148

A solution of compound 78 (63 mg, 0.18 mmol), 4 drops of concentrated hydrochloric acid, 1 mL of methanol and 1 mL of water was heated at 50 ° C for 48 h. The reaction mixture is evaporated and the residue is taken up twice with 5 ml of methanol and evaporated. The residue was taken up in 2 mL of hexane, evaporated and dried for 2 h in an Abderhalden oven at reflux with acetone to afford 69 mg (100%) of compound 148 as a white solid.

LC / MS (direct infusion, electrospray + ve, 10 mM ammonium acetate in 4: 1 water / acetonitrile) 348.20, C22H38NO2.

Table 1

compound 3α- or 3β-ΝΗ ₂ ri r ₂ r ₃ HA Hex IC ₅₀ (μΜ) Calcium (%) inhibition at 20 μΜ) 49 α ch ₃ Η Η HCl 11.0 26.8 53 α Η Η Η AcOH 17.6 -0.8 54 α Η Η Η HCl 16.1 19.8 64 α ch ₃ F Η HCl 14.4 6.7 69 α Η CO2CH3 Η AcOH 21.2 11.2 28 β ch ₃ Η Η HCl 6.8 29.0 83 β ch ₃ Η Η AcOH 7.7 30.3 89 β Η Η Η AcOH 11.5 15.0 95 β ch ₃ F Η HCl 13.5 11.2 78 α Η Η ch ₃ - 20.1 5.3 79 α Η Η ch ₃ HCl 18.4 9.1 80 α Η Η ch ₃ AcOH 10.7 7.4

Table 2

? H

compound 3a- or 3β-ΝΗ ₂ ri r ₂ HA Hex IC50 (ΜΜ) Calcium (% inhibition at 20 μΜ) 70 and 0 0 AcOH ND 7.0 72 and OH H AcOH ND 10.3 96 β CH3CH2 H HCl 7.8 47.9 97 and CH3CH2 H HCl 9.3 43.0

Table 3

compound 3a- or 3β-ΝΗ ₂ R1 r ₂ r ₃ Hex IC ₅₀ (μΜ) Calcium (% inhibition at 20 μΜ) 131 and H H ch ₃ co 14.1 9.9 132 and H H c ₆ h ₅ co 10,0 19.6 133 and H H (CH _{3) 2} CHSO ₂ 15.9 13.8 134 and H H C6H5SO2 16.7 25.1 135 β CH ₃ H ^H 0 ^{H N} h <1 15.1 37.3 137 and H H CgHsNHCO 14.8 13.0 138 and H H CH ₃ (CH ₂ ) ₂ NHCO 15.0 9.0

Table 4

compound 3α- or 3β-ΝΗ ₂ Rf r ₂ r ₃ R4 Hex IC ₅₀ (μΜ) Calcium (%) inhibition at 20 μΜ) 139 β ch ₃ ch ₃ H H 10.3 26,3 140 α ch ₃ ch ₃ H 3- (CH ₃ ) C ₆ H ₄ CH ₂ 10.2 21.5 141 α ch ₃ ch ₃ H 3,4- (F) ₂ C ₆ H ₃ CH ₂ 9.4 36.3 142 α ch ₃ ch ₃ H 3.4- (CH ₃ O) ₂ C ₆ H ₃ CH ₂ 15.9 13.8 143 β ch ₃ ch ₃ ch ₂ H H 8.6 22.6 144 α and β ch ₃ ch ₃ ch ₂ H c ₆ h ₅ 6.9 13.0 145 α and β ch ₃ ch ₃ ch ₂ H 3- (CF ₃ ) C ₆ H ₄ 22.0 -0.2 146 α ch ₃ ch ₃ ch ₂ H c ₆ h ₅ ch ₂ 9.0 41.7 147 β ch ₃ ch ₃ ch ₂ H c ₆ h ₅ ch ₂ 18.1 36.3 148 α ch ₃ ch ₃ ch ₃ H 29.2 6.6

Table 5

compound R HA Hex IC50 (μΜ) Calcium (%) inhibition at 20 μΜ) 110 , ABOUT' /. -. AcOH 10.9 39.7 111 about HCl 9.3 53.1 113 ch ₃ nh HCl 9.8 45.0 114 CH ₃ (CH ₂ ) ₂ NH HCl 10.2 37.9 115 CH ₃ (CH ₂ ) ₄ NH HCl 11.1 40.0 116 C5H9NH AcOH 9.6 60.9 117 CeHnNH AcOH 6.6 42.2 118 0 * AcOH 11.0 41.8 119 CH ₃ (CH ₂ ) ₂ NH (CH ₂ ) ₂ NH 2 HCl 6.1 54.5 120 (CH ₃ ) ₂ N (CH ₂ ) ₂ NH 2 HCl 9.1 44.0 121 H-H AcOH 14.4 42.4 122 HOCH ₂ CH ₂ NH HCl 12.3 31.4 123 HOCH ₂ (CH ₂ ) ₄ NH HCl 16.7 12.8 124 boy ₂ CH ₂ NHCH ₂ CH ₂ NH 2 HCl ND 21.0 125 3- (CH ₃ ) C ₆ H ₄ NH ₂ HCl 39.4 5.7 126 4- (HO) C ₆ H ₄ NH HCl 9.7 45.0 127 4- (H ₂ NSO ₂ ) C ₆ H ₄ NH HCl 10.1 29.4

100

compound R HA Hex IC ₅₀ (μΜ) Calcium (%) inhibition at 20 μΜ) 128 ΓΡ® Sr AcOH 10.5 41.0 129 / X ™ 2AcOH ON THE 13.8

Table 6

compound R HA Hex IC50 (μΜ) Calcium (% inhibition at 20 μΜ) 100 4 - ((CH ₃ ) ₂ CH) C ₆ H 4 AcOH 18.2 33.2 101 2- (F) C ₆ H ₄ AcOH 10.3 16.3 102 3- (CF ₃ ) C ₆ H ₄ AcOH 10,0 9.5 103 2- (CH ₃ O) C ₆ H ₄ AcOH 11.1 23.7 104 4- (CF ₃ O) C ₆ H ₄ AcOH 9.5 30.3 105 3- (C ₆ H ₅ O) C ₆ H ₄ AcOH 22.5 10.2 106 3- (NO ₂ ) C ₆ H ₄ - 8.8 8.1 107 3-C5H4N 2 HCl 17.2 11.6

101

Examples of use

Example A

Effect of selected compounds on allergen-induced pulmonary inflammation

The ability of the compound to suppress allergen-induced accumulation of inflammatory cells, such as eosinophils and neutrophils, in the lavage fluid of sensitized animals indicates the anti-asthmatic action of the compound. This model system is particularly useful for evaluating the effects of a test substance in the treatment of late stage asthma response when pulmonary inflammation and second stage bronchoconstriction occur. This test is carried out as follows.

Brown Norway rats are sensitized to ovalbumin by a single intraperitoneal injection of 1 mg ovalbumin adsorbed to 100 mg aluminum hydroxide in 1 ml sterile saline (control rats receive sterile saline only) v1. day and left for sensitization until day 21. Compounds are administered orally daily for three days before exercise (days 19, 20, 21) and one day after exercise (day 22), the third dose is administered 2 hours for exercise and the fourth dose 24 hours after exercise (volume = 300 μΙ / dose ). Rats are challenged with 0.5% ovalbumin in saline from a Devillbis nebulizer for 60 min on day 21.

h after exercise, the animals are sacrificed by an overdose of intraperitoneally administered pentobarbital sodium and lung lavage is performed with 2 x 7 ml of cold phosphate buffered saline. The resulting rinse liquid is placed on ice. The bronchoalveolar lavage fluid is centrifuged and the supernatant discarded. The pellet is resuspended in phosphate buffered saline at 4 ° C. Centrifuged cells are prepared and stained for differentiation and counting of cell types.

The protective effects of various test compounds on allergen-induced pulmonary inflammation include Tables 7 and 8. The dose-response of selected compounds is shown in Table 9. The test substance is administered in a vehicle of 300 μΙ corn oil (Tables 7 and 8) or water (Table 9). Control animals received 300 μΙ of corn oil or drug-free water alone. The values in Tables 7, 8 and 9 represent the percent inhibition of leukocyte accumulation relative to control animals. A negative score in Table 7, 8 or 9 indicates an exacerbation phenomenon above the values

102 in control animals.

Table 7

Effect of test substances (5 mg / kg / day for 4 days, orally) on ovalbumin-induced inflammatory cell accumulation in the lung lavage of sensitized rats Brown Norway

compound % inhibition eosinophils % inhibition neutrophil % inhibition cell 83 67 34 38 97 40 52 56 96 -9 40 19 64 62 70 81 89 57 60 64 28 85 87 124 53 24 57 13 95 14 45 30 49 52 36 73 135 20 58 107

103

Table 8

Effect of selected substances (1 mg / kg / day for 4 days, orally) on ovalbumin-induced inflammatory cell accumulation in the lung lavage of sensitized rats Brown Norway

analog % inhibition eosinophils % inhibition neutrophil % inhibition cell 142 -131 -1 -42 54 -82 -19 -7 107 -122 23 -28 124 -55 -76 -26 129 -296 -114 -71 146 8 24 -27 147 40 58 36 131 16 57 21 138 -52 35 33 133 -43 40 28

Table 9

Dose-dependent effect of selected compounds (repeated dosing, 4 days, 4 times daily orally) on ovalbumin-induced accumulation of inflammatory cells in lung lavage fluid in sensitized rats Brown Norway

analog % inhibition eosinophils % inhibition neutrophil % inhibition cell mg / kg 1 0.3 0.1 0.03 1 0.3 0.1 0.03 1 0.3 0.1 0.03 89 71 80 35 - 66 72 25 - 6 70 2 - 28 60 71 29 5 41 66 -10 -26 70 85 50 -5 139 42 41 46 - 63 62 42 - 56 34 19 - 143 48 29 28 - 64 25 57 - -22 -37 -43

104

Example B

Irritant-induced effect of compound 83 on ear swelling in mice

Mice are clearly identified by placing the mark on the tail. Mice receive orally 15 mg / kg of test substance in 100 μΙ 45% β-cyclodextrin in saline. Mice are rapidly anesthetized with 2% halothane and 2 µg of phorbol-12-myristate-13-acetate in 25 μΙ acetone are administered to the outer and inner surfaces of the left ear of the mouse. Acetone is applied to the right ear of the mouse in the same manner and serves as a control (vehicle). Control animals are treated in the same manner but without test substance. After 3 h, mice are sacrificed by cervical dislocation and standard biopsy samples are taken from the ears and weighed to the nearest tenth of a mg. Data are analyzed from the difference between each left and right ear and then by calculating% swelling inhibition (((mean Rx / mean with irritation)) x 100) -100.

The compounds of the invention show protective effects against ear swelling in an irritant-induced mouse. For example, compound 83 suppresses ear irritation induced by irritant in mice by 38% compared to control animals.

Example C

Effect of compounds on the release of hexosaminidase from the rat mast cell line (RBL2H3)

The antiallergic effects of a compound of the invention are evaluated by measuring its effect on antigen-induced hexosaminidase secretion from passively sensitized rat mast cells. The ability of the test substance to inhibit the release of mast cell granules, e.g. histamine and hexosaminidase, indicates the antiallergic and / or antiasthmatic activity of the compound. Hexosaminidase is released from most mast cell analogs by histamine and other mediators during antigen loading. The test is carried out as follows.

RBL-2H3 cells are grown and passively sensitized for 1 h at 37 ° C to dinitrophenol (DNP) using human anti-DNP antibody (IgE). Cells are incubated with the test substance for 30 min at 37 ° C and stimulated with 0.5 µg / ml DNP-HSA (antigen) for 30 min. Aliquots of the supernatant are removed and used for measurement

105 hexosaminidase released during antigen loading. The amount of hexosaminidase present in the supernatant is determined by colorimetric monitoring of the enzyme metabolism of p-nitrophenyl-N-acetyl-β-D-glucosaminide (p-NAG) over 60 min at 405 nm. The effect of each test substance is determined as% of the antigen-induced response (minus baseline) obtained in the presence of dimethylsulfoxide alone. These values are used to determine the degree of inhibition of antigen-induced release of hexosaminidase from cells.

The compounds of the invention demonstrate the ability to inhibit hexosaminidase release in response to antigen stimulation. Compounds are tested at concentrations of 0.3, 3, 10 and 30 μΜ and IC ₅₀ is calculated. These data are summarized in Tables 1 to 6. For example, compound 119 at a concentration of 6.1 µΜ inhibits the release of hexosaminidase by 50% due to the response to the antigen challenge.

Example D

Metabolic stability of selected compounds in human S9 fractions

The therapeutic efficacy of a test substance may be directly related to its metabolic stability in vivo. Most of the known drugs on the market are metabolised by a group of enzymes known as P450 enzymes. The human liver S9 fraction contains all P450 enzymes and cytoplasmic enzymes may also be involved in the metabolism of new chemicals. In vitro metabolism using human S9 fractions is a standard assay for evaluating the relative metabolic stability of novel compounds. This test is performed as follows.

The reagents are thawed on ice and combined to prepare Master Mix ”as follows: potassium phosphate (100 mM, pH 7), G6P (0.25 mM), G6PDH (2 units / ml), NADPH (1 mM), UDP (0.25 mM), APPS (0.25 mM) and S9 fraction (2 mg / ml). The volume of 498 μΙ of the Master mix is divided into microcentrifuge tubes. Place a volume of 2 μΙ of a 2.5 mM test compound solution (final concentration 10 μΜ) in the center of the cap of the appropriate tube and gently cap the caps to avoid mixing. A synchronous combination of the test substance and the Master mix is achieved by inverting the tubes 10 x at the same time and incubating at 37 ° C and shaking at the appropriate incubation time at 150 min * ¹ . All 0 time samples were mixed individually as incubation continued

106 is inverted three times, and then 500 μΙ of ice-cold acetonitrile is immediately added and inverted three times to stop the reaction. Immediately after each 15 minute and 30 minute incubation, ice-cold acetonitrile is added to each tube and the tubes are mixed by inverting three times. All sample tubes are incubated at -80 ° C for at least 15 min, allowed to thaw, and inverted again. Aliquots of 650 μΙ are transferred to Chromatographic Specialties tube microfiltration (0.2 .mu.m nylon membrane, C18505), and centrifuged at a speed of 13,000 rpm for 48 s ^1. Sample filtrates are stored at -80 ° C.

The sample filtrates are analyzed by liquid chromatography and mass spectrometry and the remaining percentage is calculated after incubation for 15 and 30 min relative to incubation at minute 0.

The metabolic stability of the various compounds is summarized in Table 10 as the remaining percentage after incubation for 15 or 30 min with the human liver fraction S9.

107

Table 10

Metabolic stability of selected compounds after incubation for 15 and 30 min with human liver fraction S9 as a percentage remaining from baseline after incubation time of 0 min.

compound % remaining after 15 min % remaining after 30 min 83 92 ± 19 91 ± 21 97 94 ± 10 89 ± 12 96 92 + 19 94 ± 21 89 100 ± 17 98 ± 16 28 100 ± 6 104 ± 12 49 93 ± 6 85 ± 8 64 104 ± 4 88 ± 5 139 109 + 26 100 + 10 143 100 ± 3 99 ± 10 146 82 + 6 63 ± 9 107 84 ± 5 62 ± 6 142 80 + 11 55 ± 2 69 73 ± 24 63 ± 6 104 53 ± 10 56 ± 18 141 100 + 9 26 + 4 134 85 ± 54 36 ± 15 137 93 + 31 71 + 17 138 71 ± 4 65 ± 65 132 60 + 17 40 + 9 79 67 ± 6 61 ± 14

Example E

Solubility of selected compounds in physiologically compatible compositions

The compounds of the invention exhibit good water solubility. For example

108 compound 83 has a water solubility of 225 mg / ml. Compound 83 substituted with hydroxyl on carbon C3 has a water solubility of less than 60 µg / ml. Compounds 28a have a solubility of about 30 mg / ml at room temperature, which can be significantly increased by heating. This unexpected finding shows that these 3-amino compounds can be readily formulated into therapeutic compositions.

Example F

Effect of selected test substances on the antigen-induced calcium ion flux

Increasing the cytoplasmic concentration of calcium ions is a common and fundamental event following activation of many types of cell surface receptors. The increase in intracellular calcium occurring after agonist activation of inositol lipid hydrolysis results from the release of calcium from the endoplasmic reticulum and the influx of calcium through the plasma membrane. Increases in calcium concentration in the cytosol are involved in many important cellular responses in the inflammatory process including adhesion, motility, gene expression, proliferation and degranulation. Changes in intracellular calcium concentrations may affect both short and long term cellular responses. The following is a test method to evaluate the effect of a test substance on the calcium flow.

Jurkat clone E6.1 cells grown in RPMI supplemented with 10% FBS and 2 mM L-Glutamine are transferred to 50mL conical tubes and centrifuged for 5 minutes at a speed of 900 rpm for ¹ form a cell pellet. The resulting supernatants were discarded and each pellet was washed with 10 ml HBSS. Cell suspensions were collected and centrifuged for 5 minutes at a speed of 900 rpm for ¹ form a cell pellet. The resulting supernatant was discarded and the pellet was resuspended in HBSS at a cell concentration of 1 x 10 ⁷ cells / ml. The cell suspension is transferred to a 20 mm petri dish and incubated at 37 ° C in the presence of 5% carbon dioxide for 20 min.

One volume The 2AM is mixed with one volume of Plutonic F127 detergent. Cells are labeled with 4 µl of probe solution per ml of cell suspension. The petri dish is wrapped in aluminum foil to protect it from light and placed on a shaker for 30 minutes at room temperature.

The following steps are carried out in a laminar flow fume hood

109 fluorescent lights off. The petri dish is removed from the shaker and the labeled cell suspension is transferred to a 15 ml conical tube and washed twice with HBSS as above. The cell pellet is resuspended in 12 ml HBSS and left in foil for 30 min at room temperature. An aliquot of labeled cell suspension (100 μΙ / well) is transferred to an opaque white Dynex 96 tissue culture dish. Add 50 μΙ of each test sample to the appropriate wells and incubate for 10 min at 37 ° C in a plate reader. Wallac 1320 Wind ™ (test samples are prepared in 20 mM HBSS, final concentration 20 μ 20). The selected compounds are tested for dose-related activity. Test samples and activator (anti-CD3 mAb with a final concentration of 4 pg / ml, PharMingen) are added differently (50 μΙ) so that the minimum time to reach the first data point after stimulation is about 30 s. The response of the calcium ion influx to the anti-CD3 mAb is measured as an endpoint assay ”. The entire plate is read within 100 s at a 1 s / well movement.

The dish is read prior to the addition of anti-CD3 to monitor the non-specific effect of the samples / drugs. Fluorescent emission is measured at a wavelength of 510 nm with an excitation between 340 and 380 nm every second using a pair of excitation / emission filters. Data for these two wavelengths are provided as a ratio of the values for the excitation wavelengths. This ratio is independent of intracellular dye and cell concentration and allows a true comparison between experiments.

The effect of selected compounds on calcium flux in antigen-loaded Jurkat clone E6.1 cells is summarized in Tables 1-6. For example, compound 116 provides inhibition of 60.9% at a concentration of 20 μΜ. Compound 119 has an IC 50 of less than or equal to 10 µΜ in a dose-response response. This indicates a substantial calcium flow effect that could be beneficial in any pathology of the disease in which calcium is the second major messenger or effector molecule including, but not limited to, ischemic / reperfusion injury such as stroke or myocardial infarction, inflammatory injury such as is asthma or allergy, nerve or muscle disorders such as Parkinson's disease or epilepsy, cardiac arrhythmias or hypertension.

Example G

110

Effect of selected compounds on allergen-induced changes in lung function

In asthma, the early airway response to allergen inflammation is characterized by immediate bronchoconstriction, which peaks 20-30 min after challenge and which normally resolves after about two hours. Anti-inflammatory drugs are not generally active bronchodilators and are not very effective in controlling acute asthmatic bronchoconstriction. This leads to the need for combination therapy in the treatment of bronchoconstriction and inflammation.

Cam-Hartley guinea pigs are sensitized to ovalbumin (OA) in groups of 5-6 animals by exposure to an aerosol solution of 1% OA in saline for 15 min for 2 consecutive days using a DeVilbiss nebulizer with an additional single intradermal injection of 3 µg OA in saline on day 1. Animals exhibit maximum antigen sensitivity about 14 days after initial exposure. On day 14, animals are anesthetized with ketamine (50 mg / kg intraperitoneally) and xylazine (10 mg / kg intraperitoneally), weighed and then maintained on 1% halothane delivered through the nasal cone. The left carotid artery is cannulated with a PE90 tube containing 200 units / ml heparin in saline. Tracheotomy is performed and a cannula filled with liquid (PE 160) approximately 7 cm is inserted into the esophagus. The animal is placed in a plethysmograph and the trachea is attached to a tracheal tube of stainless steel. The carotid cannula is attached to a pressure transducer to monitor blood pressure and heart rate. The guinea pig is paralyzed with pancuronium bromide (0.8 mg / kg) and ventilated at 60 Hz with a 3 ml feed volume using a Harvard small animal ventilation device.

Data are recorded for periods of 20 s at 100 Hz sampling rate on a computer physiological recording system using DIREC physiological software and analyzed with ANADAT software. Pulmonary resistance and dynamic pulmonary compliance are obtained from the value of flow and pressure signals according to the method of Von Neeguard and Wirz (1927) using an isovolumetric multipoint regression analysis model. (Ludwig, Robatto, et al., 1991), and calculated as absolute changes in lung resistance (R _Ll cm _H2O / ml / s) or lung compliance (C dyn, ml / cm / HZO). Volume and pressure signals are calibrated before each set of experiments using standard methods.

Several lung function measurements are obtained over a period of 5 to 10 min to ensure a stable baseline and then the animals are loaded with OA (2% in

Saline) administered over 6 breaths as a nebulized aerosol at a flow rate of 5 L / min. Pulmonary and cardiovascular function are continuously monitored during the experiment, although data is collected at given time points after antigen challenge (10 s, 1, 2, 3, 4, 5, 10, 20 and 30 min).

Test substances are administered under mild halothane anesthesia by oral sodium (0.1 to 1.0 mg / kg / day, daily) in 300 μΙ polyethylene glycol-200 for 4 days prior to loading with a final dose administered 2 hours prior to antigen loading.

The protective effects of selected test compounds against allergen-induced bronchoconstriction are shown in Figures 4 and 5. The duration of the activity of compound 89 is shown in Figures 6 and 7. Data are shown as mean ± standard deviation of mean. Inhibition of bronchoconstriction by test compounds would be beneficial in any disease in which acute smooth muscle constriction is a response to inflammation by an allergen, such as asthma or allergy.

All publications and patents cited in this specification are incorporated herein by reference, as if each individual publication or patent application were specifically and individually incorporated by reference. For example, a book in the Comprehensive Organic Transformations series, A Guide to Functional Group Preparations,

2nd Edition, Richard C. Larock, John Wiley &amp; Sons, Inc., 1999, and in particular the references cited herein are hereby incorporated by reference for all purposes.

It will be apparent from the foregoing that, although specific embodiments of the invention are described herein by way of illustration, various modifications may be made without departing from the spirit and scope of the invention.

112

Claims (10)

PATENT CLAIMS A compound of the general formula And R ^3, and pharmaceutically salts, solvates, stereoisomers, and prodrugs thereof, in isolation or in mixture, where independently in each case R ¹ and R ² are selected from hydrogen, oxygen to form nitro or oxime, amino, -SO ₃ -R, and organic groups of 1 to 30 carbon atoms optionally containing 1 to 6 heteroatoms selected from nitrogen atoms , oxygen, phosphorus, silicon and sulfur, where R ² may be a direct bond to atom number 3 or the R ¹ and R ² groups, together with the nitrogen atom to which they are both attached, may form a heterocyclic structure which may be part of an organic group having 1 to 30 carbon atoms and optionally may contain 1 to 6 heteroatoms selected from nitrogen, oxygen and silicon, or R ¹ may be a chain of 2 or 3 atoms attached to atom number 2 so that -NR ¹ forms part of the fused bicyclic structure of ring A . 113 R ³ and R ⁴ are selected from direct bonds to atoms 6 and 7, respectively, in such a way that carbonyl groups, a hydrogen atom or a protecting group are obtained, such that R ³ and / or R ⁴ is part of a hydroxyl or carbonyl protecting group, atoms the numbers 1 through 17 are in each case carbon atoms, wherein the carbon atoms designated by 1, 2, 4, 11, 12, 15, 16 and 17 can be independently substituted with (a) one of = O, = C (R ⁵ ) ( R ⁵ ), = C = C (R ⁵ ) (R ⁵ ), -C (R ⁵ ) (R ⁵ ) (C (R ⁵ ) (R ⁵ )) n - and - (O (C (R ⁵ )) (R ⁵ ) n O) -, wherein n is from 1 to about 6, or (b) two of the following groups are independently selected from -X, -N (R ¹ ) (R ² ), -R ⁵ and -OR ⁶ , and the carbon atoms designated 5, 8, 9, 10, 13 and 14 may be independently substituted with one of -X, -R ⁵ , -N (R ¹ ) (R ² ) or -OR ⁶ , in addition to the -OR ³ and -OR ⁴ groups mentioned, each of carbon numbers 6 and 7 may be independently substituted with one of the substituents -X, -N (R ¹ ) (R ² ), -R ⁵ or OR ⁶ , each of rings A, B, C and D is independently fully saturated, partially saturated or fully unsaturated, R ⁵ is each independently selected from hydrogen, X and C 1-30 an organic radical which may optionally contain at least one heteroatom selected from the group consisting of boron, halogen, nitrogen, oxygen, 114 silicon and sulfur, where the two geminal groups R ⁵ can form a ring together with the carbon atom to which they both attach, R ⁶ is a hydrogen atom or a protecting group, such that -OR ⁶ is a protected hydroxyl group, wherein the vicinal -OR ⁶ groups may together form a cyclic structure that protects the vicinal hydroxyl groups and wherein the geminal -OR ⁶ groups may together form a cyclic structure that protects a carbonyl group; and X represents fluoride, chloride, bromide and iodide but does not include pregnan-6,7-diol-3-one, 3-oxime, 6,7-diacetate (6α, 7β), 3-semicarbazonylhelvolic acid, 2,4-dinitrophenylhydrazone methylhelvonate Helvolic acid, 2,4-dinitrophenylhydrazone or 2,4-dinitrophenylhydrazone methylhelvolinate. A compound according to claim 1, wherein the numbers 1 to 16 are in each case carbon atoms and the carbon atoms designated by numbers 1, 2, 4, 11, 12, 15 and 16 can be independently substituted with (a) one of = O, = C (R) ⁵ ) (R ⁵ ), = C = C (R ⁵ ) (R ⁵ ), -C (R ⁵ ) (R ⁵ ) (C (R ⁵ ) (R ⁵ )) _n and - (O (C (R ⁵ ) (R ⁵ ) (R ⁵ )) _n O) -, wherein n is from 1 to about 6, or (b) two of the following substituents independently selected from -X, -N (R ¹ ) (R ² ), -R ⁵ and -OR ⁶ and the figure 17 represents a carbon atom substituted with (b) one of = C (R ^5a ) (R ^5a ), = C = C (R ^5a ) (R ^5a) ) a 115 -C (R ^5a ) (R ^5a ) (C (R ^5a ) (R ^5a )) _n -, wherein n is from 1 to about 6, or (b) two of the following substituents are independently selected -X, -N (R ¹ ) (R ² ) and -R ^5a , wherein R ^is in each case independently selected from hydrogen, X and C 1-4 organic, which may optionally contain at least one heteroatom selected from the group consisting of boron, halogen , nitrogen, silicon and sulfur, wherein the two geminal groups R ⁵ can form a ring together with the carbon atom to which they both attach. A compound according to claim 2, wherein R ^5a in all cases is independently selected from C 1-30 hydrocarbon, C 1-30 halogenated hydrocarbon, C 1-8. ₃₀ hydrogenated hydrocarbon groups, a hydrogen atom and X. The compound of claim 2, wherein R ^5a in each case is independently selected from C 1-10 hydrocarbon, C 1-10 halogenated hydrocarbon, C 1-10 hydrogenated hydrocarbon groups, a hydrogen atom, and X. A compound according to any one of claims 1 to 4, wherein R ¹ and R ² are selected from hydrogen, oxygen, such that nitro, oxime, amino, and -SO ₃ -R, and organic groups of 1 to ² are formed. 30 carbon atoms, which may optionally contain 1 to 6 heteroatoms selected from oxygen, phosphorus, silicon and sulfur atoms, where R ² may be a direct bond to the number 3 or R ¹ and R ² together with the nitrogen atom to which they are both attached to form a heterocyclic structure which may be part of an organic group having 1 to 30 carbon atoms and optionally may contain 1 to 6 heteroatoms selected from oxygen and silicon atoms or R ¹ may be a chain of 2 to 3 atoms attached to the number 2 so that -NR ¹ - forms part of the bicyclic structure condensed with ring A. A compound according to any one of claims 1 to 5, wherein 116 carbons denoted by 1, 2, 4, 11, 12, 15 and 16 are all substituted by two hydrogen atoms, unless the carbon atom considered is part of an unsaturated bond, the carbons denoted by 5, 8, 9 and 14 are all substituted by one hydrogen atom when the carbon atom considered is not part of an unsaturated bond, the carbon denoted by 10 is substituted with a methyl group and the carbon denoted by 13 is substituted with a methyl group, unless it is part of an unsaturated bond. A compound according to any one of claims 1 to 6, wherein the carbons designated 1, 2, 4, 11, 12, 15 and 16 are all substituted with two hydrogen atoms, the carbons designated 5, 8, 9 and 14 are all substituted with one atom hydrogen, the carbon of 10 is substituted with a methyl group and the carbon of 13 is substituted with a methyl group unless it is part of an unsaturated bond. The compound of claim 1, wherein R ¹ and R ² are hydrogen atoms, R ³ and R ⁴ are selected from direct bonding to atoms 6 and 7, respectively, in such a way that carbonyl groups, a hydrogen atom or a protecting group are formed, such that R ³ and / or R ⁴ is part of a hydroxyl or carbonyl protecting group; to the groups -OR ³ and -OR ⁴ , each of the numbers 6 and 7 is substituted with a hydrogen atom, unless it is excluded that -OR ³ or -OR ⁴ represents a carbonyl group, carbon atoms number 1, 2, 4, 11 , 12, 15 and 16 are all substituted with two hydrogen atoms, unless the carbon atom considered is part of an unsaturated bond, The 117 carbon atoms designated by 5, 8, 9 and 14 are all substituted by one hydrogen atom, unless the carbon atom considered is part of an unsaturated bond, the carbon atom designated by 10 is substituted by a methyl group, the carbon atom marked by 13 is substituted by a methyl group not part of an unsaturated bond, the carbon atom marked with 17 is substituted by one of = O, = C (R ⁵ ) (R ⁵ ), = C = C (R ⁵ ) (R ⁵ ), -C (R ⁵ ) ⁵ ) (R ⁵ ) (C (R ⁵ ) (R ⁵ )) to - (O (C (R ⁵ ) (R ⁵ )) n O) -, wherein n is from 1 to about 6, or (b) two of of the following groups, which are independently selected from -X, -N (R ¹ ) (R ² ), -R ⁵ and -OR ⁶ , each of rings A, B, C and D independently is fully saturated, partially saturated or fully unsaturated . R ⁵ in all cases is selected from hydrogen, X and CKo an organic radical which may optionally contain at least one heteroatom selected from the group consisting of boron, halogen, nitrogen, oxygen, silicon and sulfur atoms, wherein the two geminal groups R ⁵ may form a ring together with the carbon atom to which they both join, R ⁶ is a hydrogen atom or a protecting group, so -OR ⁶ is a protected hydroxyl group, wherein the vicinal -OR ⁶ groups may together form a cyclic structure that protects the vicinal hydroxyl groups and wherein the geminal -OR ⁶ groups may form a cyclic structure that protects carbonyl group and X is fluoride, chloride, bromide and iodide. A compound according to claim 8, wherein R ¹ and R ² are hydrogen atoms, R ³ and R ⁴ are selected from hydrogen and protecting groups such that R ³ and / or R ⁴ is part of a hydroxyl protecting group, 118 carbon atoms marked with 1, 2, 4, 11, 12, 15 and 16 are all substituted with two hydrogen atoms, carbon atoms marked with 5, 8, 9 and 14 are all substituted with one hydrogen atom, carbon atom marked with 10 is substituted methyl, the carbon atom of 13 is substituted with a methyl group, unless it is part of an unsaturated bond, the carbon atom of 17 is substituted with one of = C (R ⁵ ) (R ⁵ ) and = C = C (R) ⁽⁵ ) (R ⁵ ) or (b) two of the following groups independently selected from -X, -N (R ¹ ) (R ² ) and -R ⁵ , each of rings A, B, C and D being independently fully saturated or partially saturated, R ⁵ is independently selected from the group consisting of hydrogen, X and Cv ₃₀ hydrocarbon, halogenated hydrocarbon and hydrogenhalogenated hydrocarbon group; and X is fluoride, chloride, bromide and iodide. A compound according to claim 9, wherein R ¹ and R ² are hydrogen atoms, R ³ and R ⁴ are selected from hydrogen and protecting groups such that R ³ and / or R ⁴ is part of a hydroxyl protecting group, the carbon atoms designated by 1, 2, 4, 11, 12, 15 and 16 are all substituted two hydrogen atoms, the carbon atoms designated by 5, 8, 9 and 14 are all substituted by one hydrogen atom, the carbon atom marked by 10 is substituted by methyl, the carbon atom marked by numeral 13 is substituted by methyl, unless it is part of an unsaturated bond, 119 the carbon atom marked with 17 is substituted with (a) one = C (R ⁵ ) (R ⁵ ) or (b) two -R ⁵ , each of rings A, B, C and D independently being fully saturated or partially saturated, and Each R ⁵ is independently selected from H and hydrocarbyl CI.I _0.